Interspecific Grafting of Tomato (Solanum lycopersicum

Interspecific Grafting of Tomato (Solanum lycopersicum) onto Wild Eggplant (Solanum torvum) for Increased Environmental Tolerances

A THESIS

SUBMITTED TO THE FACULTY OF

UNIVERSITY OF MINNESOTA

BY

ANDREW J PETRAN

IN PARTIAL FULFILLMENT OF THE REQUIREMENTS

FOR THE DEGREE OF

MASTER OF SCIENCE

Emily Hoover, Adviser

September, 2013

© Andrew Joseph Petran 2013

i

Table of Contents

List of Tables………………………………………………………………………………………………..iii

List of Figures………………………………………………………………………………………………iv

CHAPTER 1

Introduction and Overview…………………………………………………………………………………1

Vegetable Production in the US Virgin Islands………………………………………………………….2

Vegetable Grafting………………………………………………………………………………………….6

Techniques………………………………………………………………………………………………….7

Disease Resistance………………………………………………………………………………………..8

Environmental Tolerance………………………………………………………………………………….9

Identification of Quantitative Trait Loci (QTLs) for Rootstock Resistance…………………………..13

Yield Differences………………………………………………………………………………………….13

Interspecific Grafting……………………………………………………………………………………...15

Interspecific Grafting in Tomatoes and Potential for use in USVI……………………………………16

Using Experiential Education to Spread Farming Techniques……………………………………….18

CHAPTER 2

Compatibility of Solanum torvum as a Rootstock in Interspecific Tomato Grafting………………..21

Summary.………………………………………………………………………………………………….22

Introduction………………………………………………………………………………………………...22

Materials and Methods…………………………………………………………………………………...26

Results……………………………………………………………………………………………………..28

Discussion…………………………………………………………………………………………………28

Literature Cited……………………………………………………………………………………………31

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CHAPTER 3

Utilization of Interspecific Grafting for Increased Flood and Drought Resistance in Tomatoes….35

Summary.………………………………………………………………………………………………….36

Introduction………………………………………………………………………………………………...36


Results……………………………………………………………………………………………………..44

ANOVA- Plant Height and Internode Length…………………………………………………………..44

Linear Regression Analysis……………………………………………………………………………...45

Discussion…………………………………………………………………………………………………45

Optimal Conditions………………………………………………………………………………………..45

Flood Conditions…………………………………………………………………………………………..47

Drought Conditions……………………………………………………………………………………….48


CHAPTER 4

Utilization of Interspecific Grafting for Increased Heat Tolerance in Tomatoes……………………63

Summary…………………………………………………………………………………………….…….64

Introduction………………………………………………………………………………………………...64


Results……………………………………………………………………………………………………..70

Discussion…………………………………………………………………………………………………71


Comprehensive Bibliography…………………………………………………………………………….79

APPENDIX

Data Tables for ANOVA and Regression Analysis……………………………………………………87

iii

List of Tables

Table 1. Scion and rootstock combinations analyzed for grafting compatibility……………………34

Table 2. Average days to fusion and survival (%) among all ‘Celebrity’ rootstock genotypes. Letters denote statistical differences (p<0.05) within rows…………………………………………...34

Table 3. Average days to fusion and survival (%) among all 3212 rootstock genotypes. Letters denote statistical differences (p<0.05) within rows……………………………………………………34

Table 1. 8 different scion and rootstock combinations were analyzed for morphological and physiological responses to different environmental pressures………………………...…………….55

Table 1. Scion and rootstock combinations were analyzed for morphological and physiological responses to different environmental pressures………………………………………………………77

Table 2. Average final plant height of all genotypes in each environmental condition. Letters denote statistical differences (p<0.05) within rows. Stars denote statistical differences (p<0.05) within columns…………………………………………………………………………………………….77

Table 3. Average final internode length of all genotypes in each environmental condition. Stars denote statistical significance (p<0.05) within columns………………………………………………77

Table 4. Average Stomatal Resistance of all genotypes in each environmental condition. Letters denote statistical differences (p<0.05) within rows. Stars denote statistical significance (p<0.05) within columns…………………………………………………………………………………………….78

Table A1: Rootstock Compatibility Trials………………………………………………………………88

Table A2: Moisture Condition Trials…………………………………………………………………….91

Table A3: Moisture Trial Final Day Calculations…………………………………………………….100

Table A4: Heat Condition Trials……………………………………………………………………….102

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List of Figures

Figure 1. Boxplot of plant height (cm) of all genotypes in each treatment, day 25. Star indicates a significant difference (p<0.05) from all related scion genotypes in that condition…………….…...56

Figure 2. Boxplot of internode length (cm) of all genotypes in each treatment, day 25… Star indicates a significant difference (p<0.05) from all related scion genotypes in that condition..…..57

Figure 3. Boxplot of stomatal resistance (mmol/m2/s) of all genotypes in each condition, day 25…………………………………………………………………………………………………...………58

Figure 4. Scatter plot showing the correlation of dry weight (g) to leaf area (cm2) of all treatments………………………………………………………………………………………………….59

Photograph 1. Randomized selection of each genotype in optimal conditions, Trial 1, Day 22...60

Photograph 2. Randomized selection of each genotype in flooded conditions, Trial 1, Day 22. Star placed above CelebrityxS. torvum to indicate significant difference in plant height and internode length…………………………………………………………………………………………...61

Photograph 3. Randomized selection of each genotype in drought conditions, Trial 1, Day 22..62

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CHAPTER 1

INTRODUCTION AND OVERVIEW

Andrew Petran

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Vegetable Production in the US Virgin Islands

There is a rich culture and history of vegetables grown in the tropics, such as the

US Virgin Islands (USVI) (Palada and Crossman, 1999). The USVI Agricultural

Experiment Station states that the production and marketing of fruits and vegetables

provides a primary or secondary source of income, and “many Virgin Islanders

supplement their diets with fruits, vegetables and herbs grown in home gardens” (USVI

Agricultural Experiment Station, 2000). These plants are rich sources of nutrients and,

when exported to interested markets such as the continental US, can be a viable

addition to domestic economies (Oomen and Grubben, 1978). Tropical leaf vegetables

have unique adaptations that allow them to survive in tropical climates that have widely

fluctuating amounts of rainfall throughout the year. According to Sands (1974), average

monthly rainfall on the St. Croix Island of the USVI ranges from 4.26 to 6.08 inches

August through November, and declines to a low of 1.71 inches in March and throughout

most the summer. Tropical vegetables that can survive in these variable conditions

include amaranth, eggplant (Solanum melongena), pak choi (Brassica rapa), malabar

spinach (Basella rubra), sweet potato (Ipomoea batatas) and bush okra (Abelmoschus

esculentus) (Palada and Crossman, 1999).

Staple vegetables of the continental United States, such as tomatoes (Solanum

lycopersicum), cabbage (Brassica oleracea), onions (Allium cepa) and peppers (Piper

nigrum) are incredibly popular in the USVI as well, and several attempts at domestic

production have been made (Pena and Hughes, 2007; Sands, 1974). Successful

integration of these vegetables into tropical food production systems would have

financial, cultural and nutritional benefits. The Asian Vegetable Research and

Development Center (2006) states that, concerning tropical communities, “Vegetables

are the best resource for overcoming micronutrient deficiencies and provide smallholder

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farmers with much higher income and more jobs per hectare than [native] crops”. Sands

(1974) points out that having a more diverse and fully developed agricultural system

would complement the USVI’s major industry –tourism- by offering visitors more fresh

local products that wouldn’t otherwise be available. He goes on to argue that from a

cultural standpoint, expanded agriculture would preserve the natural environment of the

USVI as well. Geographic potential of expanded agriculture exists- 14.7% of USVI land

area exists as arable and permanent cropland, compared to an 11.3% world average

(Earth Trends, 2003).

Expansion and diversification of USVI fruit and vegetable crops is limited and

constrained, however, by a complex of biological, physical and socioeconomic factors.

Crops must be able to handle a short period of extremely wet soils and harsh, rainy

conditions followed by an extended dry season while the crops are maturing. Rainfall

records at the Frederiksted USVI weather station show that less than once inch of rain

fell in the months of February, March and May five times in a twelve year span from

1959-1972, and over six inches of rain averaged in October and November (Sands,

1974). Indeed, the problem still exists today, as the University of Virgin Islands

Agricultural Experiment Station listed the short rainy season as the major limiting

constraint of USVI agricultural production (2000), and Pena and Hughes (2007) state

“environmental stress is the primary cause of crop losses worldwide, reducing average

yields for most major crops by more than 50%”. Most vegetable crops prefer cooler

temperatures and constant moist soils (Ali, 2000), and the environmental extremes

present in the USVI and other tropical areas pose a significant challenge. Vegetables, by

definition, are succulent plants, and most are comprised of more than 90% water

(AVRDC, 1990). Thus, water supply in the soil and the atmosphere has incredible

influence on the size and quality of vegetables. The dry seasons common in the USVI

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climate lead to high rates of evapotranspiration and an increased solute concentration in

the soils, causing an osmotic flow out of plant cells (Jipp et al., 1998). This lowers the

water potential within the plant, and disrupts complex cell processes such as

photosynthesis and respiration. Too much water in the rainy season poses yet another

problem. Flooded soils inhibit oxygen processes in the root zone, and waterlogging is

proven to produce endogenous ethylene that causes damage to tomatoes (Drew, 1979).

Temperature stress, in addition to water stress, reduces vegetable production,

and climatologists believe that the combined environmental stress in the tropics will only

get worse over time. Climate trend analysis in tomato growing regions by Bell et al.

(2000) show that temperatures are going to rise, and the frequency and severity of

above-optimal temperatures for vegetative growth are going to increase in the coming

decades. This will prove to be detrimental to tomatoes specifically, as their growth and

reproductive processes are strongly modified by increases in temperature alone, and

also in conjunction with other changing environmental factors like water stress (Abdalla

and Verderk, 1968). Maximum growth for most tomato plants occurs at day and night

temperatures of 25 degrees Celsius (Max et al., 2009). The average temperature in the

USVI however is 27 degrees Celsius, and average max temperatures often climb as

high as 32 degrees Celsius (Brown and Lugo, 1990). Fundamental biochemical

reactions are disrupted by high temperatures in most vegetable crops, such as a

decrease in photosynthetic capabilities, reduced fruit set and pre-anthesis temperature

stress leading to poorly developed flowers (Weis and Berry, 1988; Stevens and Rudich,

1978; Sato et al. 2002). In pepper, another solanaceous vegetable, high post-pollination

temperatures inhibited fruit set, suggesting that fertilization is sensitive to high

temperature stress (Erickson and Markhart, 2002). In summary, supra optimal

temperatures in tomato crops induce fruit set failure by bud drop, abnormal flower

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development, poor pollen production, ovule abortion and poor viability, reduced

carbohydrate availability, and reduced vegetative growth due to an inhibition of

photosynthesis (Hazra et al. 2007).

Improving agricultural infrastructure would offset some of the environmental

problems that face plant production in the USVI, but many are too expensive to be

considered and government support for agricultural sectors has been declining for the

past 50 years (McElroy et al., 1990; Sands, 1974). Due to financial constraints, the USVI

has no wheeled or crawler tractors and zero percent irrigation (EarthTrends, 2000;

Mwaijande et al., 2009). Greenhouses are used to change the microclimate around

plants to create favorable growth conditions, but even if the USVI did improve its

agricultural infrastructure with modern greenhouses, outside climatic pressures would

still prove troublesome. Tropical greenhouses are often naturally ventilated to reduce

energy costs (Max et al., 2009). This method of passive cooling relies on prevailing

atmospheric conditions, especially wind, and ventilation openings need to have a large

surface area in order to effectively cool the greenhouse (Kittas, 1996). According to

Harmanto et al. (1996), ventilation openings in these types of greenhouses in lowland

tropical areas also need to be covered with insect screens that decrease wind velocities

and air exchange. This creates a less favorable microclimate within the greenhouse

area, and would negatively affect plant growth (Harmanto et al., 1996). Thus, the

expense of installing and maintaining greenhouses would not likely yield profitable

results in the USVI.

Finally, cultural and economic shifts have reduced tax incentives and rural

interest in USVI agriculture as well. Beginning in the postwar era, a decline in total farm

acreage from 44,062 to 17,785 coincided with a 400% increase in tourism and 300%

increase in hotels (McElroy and Tinsley, 1982). This increase is likely due to commercial

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tourism land being up to 1,000% more profitable per acre than land devoted to

agriculture- tourism today accounts for more than 50% of the USVI GDP, up from 10% in

1960 (BEA, 2012; McElroy et al., 1990). Thus, agriculture has experienced a decline in

favor from the native workforce, as available labor has fled from rural areas to work in

the more lucrative tourism market.

These combined factors have all but destroyed large-scale vegetable production

in the USVI. According to a 2003 report by EarthTrends, the USVI had no cereal or

solanaceous crops in major commercial production, despite its geographic potential for

expanded agriculture. The University of the Virgin Islands Agricultural Experiment (2000)

Station found that the USVI only produces five percent of its total vegetable

consumption. This has left the USVI with little to no food security. Multiple innovations in

vegetable production that are simple, affordable and accessible will be needed if these

obstacles are ever to be overcome in the USVI and tropical regions worldwide.

Vegetable Grafting

Improving environmental and climatic stress tolerance through grafting is an

approach that has been in use in East Asia during the 20th century. In this context,

grafting involves joining together two living plant parts- a rootstock and a scion- to

produce a single, living plant. While grafting is an incredibly popular technique for

increasing tomato and vegetable production in East Asia, it is almost nonexistent in the

western hemisphere. According to a project report from Ohio State University (2006), “It

is an accident of history that grafted vegetables dominate the high-tech hydroponic

greenhouse industry and are very common in subsistence production in Asia, but are

nearly absent from U.S. soil-based production systems”. The report goes on to

speculate that conventional farmers in the U.S. would shy away from grafting as an

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alternative to traditional fumigation techniques, but since conventional farming is so

limited in the USVI, it is unlikely that such a cultural anxiety would exist.

There are various techniques utilized to graft the scion onto the rootstock, and

the proper selection of technique is dependent upon the size, growth stage and

compatibility of the two plants in question.

Techniques

The three most common grafting techniques for tomato production are tube, cleft,

and approach grafts (McAvoy, 2005; Toogood, 1999). In tube grafting, a 45-degree

diagonal cut is made through the entire stem in both the rootstock and the scion. Ideally,

both cuts are made below the cotyledon, as this reduces the chance of the rootstock

suckering after the graft has healed (Bausher, 2011). The two pieces are joined together

and the graft union is covered by either a grafting clip or plastic parafilm (Toogood,

1999). Newly grafted plants are immediately brought into a ‘healing chamber’- a low light

environment with high relative humidity and a minimum of 18 degrees C at all times.

After approximately seven days the plants can be removed from the chamber. Tube

grafting is ideal for young plants with small stem diameters when only a single cut is

manageable (McAvoy, 2005). Since the plants are smaller, tube grafting allows more

plants to be put into the healing chamber at one time, but vascular contact in the graft

union of tube grafts is minimal. Vascular regeneration reestablishes the continuity of

water transport through the graft union, and a high level of vascular contact between the

scion and rootstock expedites this process (Fernandez-Garcia, 2004). Since tube

grafting has the least amount of vascular contact between the rootstock and scion

among the three techniques, the risk of graft failure in the healing chamber is high.

Cleft grafting is most commonly used on solanaceous crops (Lee and Oda,

2003). Both the rootstock and scion are grown to larger minimum stem diameters than

8

with tube grafting, when the plants are at about the 4-5 leaf stage (McAvoy, 2005).

Similar to tube grafting, the rootstock is cut off below the cotyledon, but in cleft grafting a

second, longitudinal cut is made 1.5 cm deep, about 75% the depth of the stem. The

scion is pruned to 1-3 leaves, reducing transpiration in the healing chamber, and the

lower stem is cut into a tapered wedge to place inside the depth cut of the rootstock (Lee

and Oda, 2003). Since plants that are cleft grafted are larger and have more vascular

contact at the graft union, success rates are higher than tube grafting. This success,

however, comes at the expense of longer time needed to develop the plants before

grafting, and the larger size of the plants, reducing the total number that can be grown in

limited greenhouse space.

Approach grafting is a longer, two-step process that ensures the highest

comparable survival rate, and is used often in both tomato and eggplant grafting

systems (Yamakawa, 1982). In an approach graft, a 45-degree tongue is cut into the

stem of both the scion and the rootstock without cutting off the entire plant. The two

tongues are then fitted into each other and secured with a grafting clip or parafilm, and

put into the healing chamber. While healing, the scion is still receiving water and

nutrients from its own roots, and thus risk of graft failure is low (Oda, 2006). After 3 to 4

days in the healing chamber, the scion is completely removed from its roots. Approach

grafting is slowly losing popularity among commercial growers because of the extra labor

and time involved in cutting the rootstock twice, larger space demand compared to other

methods, and a generally weaker graft union more prone to breaking or scion rooting

after transplanting (Lee et al., 2010).

Disease Resistance

Grafting is an ideal technique for vegetable production because scions with

desirable fruit-producing traits that are also susceptible to soil-borne disease or climatic

9

pressures can be grafted onto rootstock that is more resistant to these pressures. The

resulting union often results in a more productive plant (Cohen et al., 2002; Miller et al.,

2005). This solution is preferable to breeding resistant varieties of desirable plants,

which can be time consuming, expensive, technically demanding and sometimes

controversial (Sleper et al, 1991). Grafting requires no herbicide or pesticide input, and

thus would not affect the minimal agricultural infrastructure that exists in the USVI today

(EarthTrends, 2003). Grafting has primarily been used to reduce the occurrence of soil-

borne disease in non-native fruit vegetable plants, primarily tomato, pepper, eggplant

and various Cucurbitaceae (Black et al., 2003; Palada and Wu, 2008; Rivard and Louws,

2008). In tomatoes specifically, grafting onto tolerant rootstock has been used to

suppress verticillium dahlia infection since the 1960s (EarthTrends, 2003; Zhou et al.,

2009). Rivard and Louws (2008) found that ‘German Johnson’ heirloom tomatoes had

0% fusarium wilt incidence in infested soils when grafted onto resistant CRA 66 or

Hawaii 7996 tomato rootstock, compared to a 79% incidence on nongrafted controls. If

an appropriate rootstock is used, grafting can also provide tolerance to soil related

environmental stress such as drought, salinity and flooding- some of the primary

environmental challenges posed by USVI agriculture. Romero et al. (1997) grafted

melons onto a hybrid squash rootstock, and found that the grafted melons were more

salt tolerant than non-grafted controls. Despite these advantages, vegetable grafting for

disease resistance is still rare in the western hemisphere (Kubota et al., 2008), and no

peer-reviewed research exploring the potential benefits vegetable grafting in the USVI

could be found.

Environmental Tolerance

In addition to increased disease resistance, vegetable grafting has been found to

reduce the detrimental effects of abiotic stresses as well, including sub-optimal

10

temperature, moisture and salinity conditions (Schwarz et al., 2010). Temperature

stresses, both too high and too low, have been shown to lead to inhibited plant growth

and development, wilt, necrosis, and reduced overall yield of curcubit and solanaceous

crops(Ahn, 1999). Sub-optimal temperatures have been shown to reduce root growth

architecture, inhibit nutrient absorption, water absorption and translocation, disrupt

sink/source relations and slow phytohormone transport (Ali et al., 1996; Bloom et al.,

2004; Choi, 1995; Venema et al., 2008; Tachibana, 1982). Conversely, supra-optimal

temperatures have observed deleterious effects on crops as well, including growth

reduction, decrease in photosynthetic capacity accompanying increased respiration,

osmotic damages and inhibited ion uptake/transport (Wang et al., 2003). Such

conditions are of special concern to growers located in the lowland tropics such as the

USVI (Palada and Wu, 2006).

One solution to these negative responses has been to alter the immediate

growing environment of the crops by installing greenhouse systems utilizing active

protection. These systems allow growers to bring temperatures back into their optimal

threshold range for crops, but increasing energy costs has rendered this option

inefficient and untenable in many areas of the world (Schwarz et al., 2010). Indeed, a

statistical review by Winters and Martins (2004) states that territories with ‘micro-

economies’, which includes the USVI and many other Caribbean countries, are at a

73.4% electricity cost disadvantage when compared to larger, more developed

economies. The electricity rate for the USVI at the beginning of 2013 was 54.3

cents/KWh for commercial enterprises, compared to a US national average of 8.28

cents/KWh in 2011 (MCL, 2011; WAPA, 2013). Thus, it becomes apparent that lower-

input solutions must be made in these areas.

11

One explored alternative is to breed cash crops with larger optimal temperature

thresholds, instead of attempting to alter environmental temperature itself. This would

allow plants to avoid detrimental physiological responses to normally suboptimal

temperatures without active energy input. Though traditional breeding has increased

production and made cash crop yields in greenhouses more efficient, little progress has

been made on breeding plants with vastly expanded temperature tolerances (Van der

Ploeg and Heuvelink, 2005). Temperature tolerance is developmentally regulated,

growth stage specific, and involves many genes (Schwarz et al., 2010). Thus, the gene

characterization of temperature tolerance is not well known, and efforts to successfully

breed expanded temperature thresholds in plants, and tomatoes specifically, are limited

(Foolad and Lin, 2001).

A method used to expedite the slow breeding process is to utilize vegetable

grafting. High performance commercial scions can be grafted onto select rootstock with

naturally wider optimal temperature thresholds. Multiple projects have demonstrated

that tomato rootstock plants with high temperature tolerances have the ability to transfer

tolerance to scions when appropriately grafted (Bloom et al., 2004; Venema et al., 2008;

Zijlstra and Nijs, 1987). This resulted in a greater quantity of flowers, trusses and fruit

production in grafted plants when compared to non-grafted controls at temperatures

considered non-optimal for the scion. Venema et al. (2005) notes that for tomatoes,

these naturally occurring wider temperature thresholds are much more common in wild

species because of the reduced genetic diversity that is present in inbred, domesticated

tomatoes.

Moisture stress is becoming an increasingly important issue in agriculture,

especially in arid and semiarid regions of the world (Schwarz, 2010). In the USVI

specifically, environmental flood and drought stresses are known to be two of the most

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limiting factors for vegetable production (UVI Ag. Experiment Station, 2000; Sands,

1974). Drought conditions lead to reduced metabolic activity and higher plant salinity,

damaging plants and yields, while flood stresses lead to deoxygenation of soils, starving

plants of the oxygen needed as a final electron acceptor in aerobic respiration. This

slows photosynthetic rate, chlorophyll content and transpiration in the plant (Kato et al,

2001). Combined with a systemic lack of irrigation infrastructure, the USVI is left highly

vulnerable to unfavorable moisture conditions (EarthTrends, 2000).

In grafted plants, the osmotic potential of dehydrated scions is determined by the

drought tolerance of its rootstock (Sanders and Markhart, 1992). In kiwifruit, grapes and

soybeans there are proven drought-tolerant rootstocks that are available and effective in

commercial growing conditions, but little is known about the potential of grafted drought

tolerance in fruit vegetables such as tomatoes (Clearwater et al., 1994; Schwarz et al,

2010; Serraj and Sinclair, 1996). Also, the depression in photosynthetic rate, stomatal

conductance, transpiration and soluble proteins associated with flood stress has been

lessened when crops are grafted onto flood-tolerant rootstock (Kato et al., 2001). Thus,

improving water use efficiency in drought conditions and flood resistance of cash crops

by grafting onto tolerant rootstocks is an appealing low-input option.

Recently, promising research has reported that grafting vegetables can alter their

ability to filter uptake of potentially harmful organic pollutants. Otani and Seike (2007)

found that when cucumber was grafted onto various Cucurbita spp. rootstock, certain

combinations displayed a significant reduction in the uptake of aldrin, deildrin and endrin,

highly toxic organic pollutants. As with temperature, drought and flood tolerances, the

correct rootstock must be selected for optimal performance, as genetic diversity among

rootstocks vary greatly.

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Identification of Quantitative Trait Loci (QTLs) for Rootstock Resistance

Studies on the genetic basis for environmental tolerances in vegetables crops

are sparse, but Venema et al. released a review of the genetics surrounding sub-optimal

temperature tolerance in tomatoes in 2005. Through backcrossing, several genes and

QTLs have been identified that control the plastochron index, shoot turgor pressure and

root growth of tomatoes in suboptimal temperatures (Foolad and Lin, 2001; Goodstal et

al., 2005; Truco et al., 2000; Vallejos and Tanksley, 1983). Further studies are needed to

identify QTLs for environmental tolerances in tomatoes and other vegetable crops. Once

accomplished, an efficient introgression of genes conferring specific environmental

tolerances into rootstocks could be made, possibly via Marker Assisted Selection.

Schwarz (2010) however points out that before such research can be done, we must first

“…identify [the] physiological characteristics that reflect the complex underlying genetic

make-up [of environmental tolerances]”.

Yield Differences

The ultimate goal of any farmer is to adopt efficient cultural practices that

maximize profit output with minimal resource input. Thus, grafted crops with enhanced

environmental tolerances will only be utilized on a commercial scale if those tolerances

lead to higher yields, and ultimately more profit. The following section explores the effect

of vegetable grafting on increasing the yield of crops under disease, temperature and

moisture pressures.

Grafting plants to the correct rootstock for resistance to microbial pathogens will

result in higher total yields and increased profit for commercial farmers. An NCSU

research project found that grafting with Maxifort tomato rootstock increased yield in high

tunnels where disease pressure from Verticillium wilt was present (Groff, 2009). The

14

grafted tomatoes allowed for an approximate 20 percent increase in yield, representing

9.4 more tons per acre. This increased yield translated into an additional gross income

of $9,024 per high tunnel acre, or $1.88 per plant, more than offsetting the marginal

added costs of growing grafted tomatoes opposed to control (Groff, 2009). Other

diseases known to be suppressed by proper rootstock selection leading to higher yields

are Fusarium oxysporum in cucurbit and tomato crops, verticillium wilt in eggplant,

cucumber and watermelon, Ralstonia solanacearum bacterial wilt in tomato and

Phytophthora capsici in cucumber (King et al., 2008).

Explorations of grafted plants growing in suboptimal temperature conditions on

total yield have shown highly variable results. This affirms that one must select the

correct rootstock from a diverse gene pool to target specific environmental stresses.

Initial trials by Okimura et al. (1986) and Bulder et al. (1987) on Cucurbitaceae

and Solanaceae showed that different scion-rootstock combinations don’t respond with

significantly different yields in suboptimal temperatures, but subsequent trials have

identified rootstocks that lead to higher overall yields in tomato, cucumber and

watermelon (Ahn et al., 1999; Davis et al., 2008; Tachibana, 1982; Ziljstra and Nijs,

1987). Not surprisingly, all grafted combinations that show suboptimal temperature

tolerances are those with rootstocks with wide optimal temperature thresholds (Schwarz

et al, 2010).

In supraoptimal temperature conditions, research has shown that grafting

tomatoes onto heat-tolerant tomato rootstock increases vegetative growth, but with no

significant difference in yield compared to non-grafted controls (Abdelmageed and

Gruda, 2009). However, using eggplant rootstock may be more promising for

supraoptimal temperature conditions, since eggplants are more adapted to live in hot,

arid climates (Abdelmageed and Gruda, 2009; Schwarz et al., 2010). Indeed, Wang et

15

al. (2007) observed yield increases of 10% on eggplant when grafted onto heat-tolerant

eggplant rootstock. Chili pepper rootstock has been shown to be an effective option in

supraoptimal temperature conditions as well (Palada and Wu, 2008).

Many tomato cultivars and lines are susceptible to supraoptimal temperatures

that would be present in the USVI (Abdalla and Verderk, 1968). Most tomatoes grow

under optimum day/night temperatures of 25 degrees Celsius (Max et al., 2009),

however some tomato lines are tolerant well above that mark. Abdul-Baki (1991) found

that several tomato lines genetically selected for heat tolerance produced a higher yield

in high temperature conditions (38-40 degrees Celsius) than when they were grown in

normal field conditions (26-28 degrees Celsius).

Even without the presence of temperature, moisture or pathogen stress it has

been found that grafted tomatoes can still lead to increased yields compared to non-

grafted controls, especially in older heirloom cultivars (Rivard and Louws, 2008). Thus,

grafting tomatoes may be a financially viable cultural practice even in optimal production

conditions. Possible explanations for vegetable yield increase in optimal conditions

include increased water and macronutrient uptake by vigorous rootstock genotypes

(Ruiz and Romero, 1999; Yetisir and Sari, 2003). Fernandez-Garcia et al. (2002) found

that certain rootstocks can improve the stomatal conductance of tomato scions.

Interspecific Grafting

Interspecific grafting is a more recent development to help further the

environmental tolerances of vegetable crops. While grafting vegetables onto tolerant

rootstock of its own species has been proven to increase resistance to various

environmental pressures such as flood, drought, cold, heat and pathogen stress (Bloom

et al., 2004; Sanders and Markhart, 1992; Venema et al., 2008; Zijlstra and Nijs, 1987),

16

in some cases the transferred tolerance is not strong enough, or a certain desired

environmental tolerance does not yet exist within the rootstock germplasm of that

species. Interspecific grafting can help to alleviate this problem. Vegetable species with

certain environmental susceptibilities will sometimes have compatible relatives with a

natural resistance to that stress. After grafting Solanum melongena L. eggplant scions

onto a verticillium wilt resistant tomato rootstock Lydl, Liu and Zhou (2009) found a 0%

incidence of the disease on grafted eggplant, compared to a 68.3% incidence on

nongrafted controls. Allelopathic chemicals were also found in the tomato rootstock

exudates, inhibiting spore germination and mycelium growth. Davis et al. (2008) states

that watermelon grafted onto the rootstock of bottle gourd (Lagenaria siceraria) confer

significant resistance to Fusarium spp. Interspecific grafting has been used successfully

for many vegetables in the curcubit and solanaceous families (King et al., 2010).

Interspecific Grafting in Tomatoes and Potential for use in USVI

Tomatoes are one of the most lucrative cash crops worldwide, but they are

especially sensitive to excessive flooding or drought, making them difficult to produce in

tropical regions (EarthTrends, 2003; Max et al., 2009). Multiple accessions of tomato

rootstock are in use commercially to confer temperature and salinity tolerances, but to

date no tomato rootstock has significant resistance to flood conditions (Bhatt et al., 2002;

Fernandez-Garcia et al., 2003; Schwarz et al., 2010). Thus, if a compatible rootstock of

another species with drought tolerance could be found, it would help fill the gap of flood

tolerance in tomato production. Eggplant (Solanum melongena) is a highly resilient and

closely related solanaceous plant to tomato with many cultivars whose roots can survive

for several days underwater.

17

There is a history of grafting tomato scions onto eggplant rootstock to mitigate

unfavorable climatic conditions, and vice versa. Okimura et al. (1986) found that

eggplants grafted onto S. integrifolium x S. melongena rootstocks grew better at lower

temperatures (18oC to 21oC) than non-grafted plants, and Midmore et al. (1997) states

“Tomato scions grafted onto eggplant rootstock grow well and produce acceptable yields

during the rainy season.” Black et al. (2003) recommends using eggplant rootstock for

tomatoes when flooding or waterlogged soils are expected, and to select lines that are

resistant to bacterial wilt and other soil borne diseases. The AVRDC has found that

eggplant accessions EG195 and EG203 are compatible with most tomato scions and

resistant to flooding, bacterial wilt, fusarium wilt and root-knot nematode, and Chetelat

and Peterson (2003) have identified tomato accession ‘Hawaii 7998’ as broadly

compatible with distantly related solanaceous crops, such as eggplant and pepper. Pena

and Hughes also state “In addition to protection against flooding, some eggplant

genotypes are drought tolerant and eggplant rootstocks can therefore provide protection

against limited soil moisture stress.” (2007). This is likely due to eggplant being more

effective at water uptake than tomato root systems (Schwarz et al., 2010).

The identification of an ideal rootstock for interspecific grafting can be achieved

by surveying available research, or by one’s own experimental findings. When starting

an initial search for interspecific rootstock, it may be wise to begin with plants native to

the region where the crop will be cultivated. This way, the likelihood of compatible

rootstock having appropriate environmental tolerances to be transferred to the non-

native scion may be increased. If a compatible native rootstock also with a history of

successful use can be found, then probability of success is even greater.

A USVI species of wild eggplant, Solanum torvum, may be an ideal rootstock

candidate for tomatoXeggplant grafting in the USVI. As a native plant of St. Thomas and

18

St. Croix it already exhibits tolerance to the climatic pressures of tropical regions. There

is also an established history of S. torvum for use as a rootstock in S. melongena

cultivation for its resistance to a wide range of soil borne pathogens, including

Verticillium dahlia, Ralstonia solanacearum, Fusarium oxysporum and Meloidogyne spp.

root-knot nematodes (Bletsos et al., 2003; Gisbert et al., 2011; Singh and

Gopalakrishnan, 1997). Uniform production of S. torvum rootstock seedlings can be

challenging as a result of low germination rate leading to poor seedling emergence and

slow early growth (Liu and Zhou, 2009). This potential hurdle can be overcome in the

USVI, however, because of the vast wild population and cheap, easy access to seed

year-round.

Using Experiential Education to Spread Farming Techniques

Global climate change has been predicted to negatively affect agricultural

production worldwide, and it is likely that regions without developed agricultural

infrastructure such as the USVI will be most damaged by these environmental changes

(Bell et al., 2000). Thus, not only is there a need for research in low-input vegetable

production techniques in environmental stress, but also an effective methodology for

broadcasting this research to farmers. Utilizing progressive, experiential education may

be effective in this case because the specific problem is narrow, specialized and location

based. America’s most respected educational scholars have argued for experiential

education that places learners in a real-world context, engaging them in meaningful

activities that focus on a specific interest (Dewey, 1916; Parr et al., 2007). Experiential

education- the interaction of a learner with his/her environment- is thus intrinsically

connected with the plant sciences, acquainting students with their natural environment.

The use of experiential agroecology education has been suggested to efficiently spread

19

the newest production techniques to the producers themselves; especially if the

producers do not have direct access to published research (Francis et al., 2011).

Incorporating experiential education into an entire curriculum or even a single

topic has been shown to increase learner retention and promote self-inquiry. Anderson

and Piscitelli (2002) found that adding an informal learning environment to the formal

teaching of a topic increased the quality and retention of overall curriculum objectives

among students. Bauerle and Park (2012) showed that the addition of a place-based

educational field trip increased the homework scores of students in a plant biology

course at Cornell University compared to students who did not attend the trip. This

increase was even more pronounced in students taking the class who were not majoring

in plant sciences. Observed increases in retention and performance can likely be

attributed to the theories of Lewin (1951), finding that learning is a continuous cycle

when concrete experiences are involved, causing the learner to observe and reflect

about the actual experience, rising to conceptual generalizations which can then be

applied to future experiences.

Experiential techniques have already been used successfully in applied

vegetable grafting education. Dissemination of grafting techniques to local farmers and

gardeners increases grafting success rate and efficiency in the field (Heinrichs et al.,

2008). The World Vegetable Center documents seminars given in India to provide

training to staff and collaborating farmers on vegetable grafting technologies. In the

same light, USVI grafting seminars would provide training to teachers at the University

level and give direct, experiential learning to both commercial producers and interested

backyard hobbyists. It is not unlikely that these experiential seminars would provide the

same success and increase the likelihood of interspecific grafting being utilized in an

area where it could be an effective production tool. Seminars would also be particularly

20

important if using native eggplant rootstock (S. torvum) that is difficult to produce in

greenhouse conditions.

21

CHAPTER 2

COMPATIBILITY OF SOLANUM TORVUM AS A ROOTSTOCK IN INTERSPECIFIC TOMATO GRAFTING

(for submission to HortTechnology)

Andrew Petran

22

Summary

Two tomato scions (‘Celebrity’ and ‘CLN3212A’) were grafted onto wild eggplant

(Solanum torvum) rootstock to determine compatibility as a rootstock for interspecific

grafting. S. torvum was compared against ‘Maxifort’, self grafted and non-grafted control

rootstock in this experiment. Seed sown S. torvum rootstock was also compared against

rooted vegetative cuttings of S. torvum to determine if there is a difference in

compatibility based on method of rootstock propagation. Average days until graft fusion

and survival rate was taken for each genotype. Vegetative S. torvum cuttings had the

poorest grafting success rate as a rootstock (50% for both scions), while all other

rootstock genotypes had statistically similar success rates. There was no significant

difference in time to graft fusion among any grafted genotypes. High compatibility of

seed sown S. torvum suggests it’s potential use as an interspecific grafting rootstock

with cultivated tomato.

Introduction

Plant grafting has been utilized in agriculture since the first millennium BCE

(Mudge et al., 2009). The process involves joining together two parts (a rootstock and

scion) from different plants to form a single, living plant. Over most of it’s history, grafting

was centered around woody perennials as a method to asexually propagate species that

did not root well from vegetative cuttings, but starting in the 20th century grafting began

to be used extensively on annual vegetable crops as well (AVRDC, 1990). Especially

popular in East Asia, vegetable grafting allows a grower to combine a scion with

desirable fruit producing traits with a rootstock that is resistant to a multitude of

environmental pressures, such as climate and pathogen stress. The resulting union

often results in higher yields (Cohen et al., 2002; Miller et al., 2005).

23

Grafting vegetable scions onto a rootstock of its own species is common

because intraspecific compatibility is often very high (Black et al., 2003; Palada and Wu,

2008; Rivard and Louws, 2008). Intraspecific grafting has been shown to increase

resistance to various environmental pressures such as flood, drought, cold, heat and

pathogen stress, however in some cases the transferred tolerance is not strong enough,

or a certain desired environmental tolerance does not yet exist within the rootstock

germplasm of that species (Bloom et al., 2004; Sanders and Markhart, 1992; Venema et

al., 2008; Zijlstra and Nijs, 1987). Intraspecific grafting would not be a viable cultural

practice in these unique circumstances, but grafting is not always limited to intraspecific

interactions. Vegetables with certain environmental susceptibilities will sometimes have

graft-compatible relatives within the same genus that possess a natural resistance to

that stress. Thus, interspecific grafting can be used to broaden rootstock diversity when

environmental pressures surpass the advantages that can be provided by intraspecific

grafting alone.

While not as common as intraspecific grafting, the successful use of interspecific

grafting in vegetable production is well documented (King et al., 2010). After grafting

Solanum melongena L. eggplant scions onto a verticillium wilt resistant tomato (Solanum

lycopersicum) rootstock Lydl, Liu and Zhou (2009) found a 0% incidence of the disease

on grafted eggplant, compared to a 68.3% incidence on nongrafted controls. Allelopathic

chemicals were also found in the tomato rootstock exudates, inhibiting spore

germination and mycelium growth. Davis et al. (2008) states that watermelon grafted

onto the rootstock of bottle gourd (Lagenaria siceraria) confer significant resistance to

Fusarium spp.

To identify a useful rootstock for interspecific grafting, first a relative with unique

environmental resistances must be found, and then tested for rootstock compatibility.

24

Interspecific grafting compatibility is difficult to predict because the degree of taxonomic

affinity necessary for compatibility varies widely across different taxa (Mudge et al.,

2009). Four potential mechanisms of interspecific incompatibility are identified by

Andrews and Marquez (1993): cellular recognition, wounding response, plant growth

regulators, and incompatibility toxins. Since prediction is difficult, individual grafting trials

must assess compatibility.

Tomatoes are lucrative cash crops with worldwide appeal, but they are sensitive

to excessive flooding or drought, making them difficult to produce in tropical regions

(EarthTrends, 2003; Max et al., 2009). Multiple accessions of tomato rootstock are in

use commercially to confer temperature and salinity tolerances, but to date no tomato

rootstock has significant resistance to flood conditions (Bhatt et al., 2002; Fernandez-

Garcia et al., 2003; Schwarz et al., 2010). Thus, the identification of a flood tolerant

rootsotck would help fill the gap of flood tolerance in tomato production. One potential

candidate is eggplant (Solanum melongena), a highly resilient and closely related

solanaceous plant to tomato with many cultivars whose roots can survive for several

days underwater (reference for this?).

Tomato/eggplant interspecific grafting has a history of successful use for

conferring environmental tolerances to fruit producing scions. Okimura et al. (1986)

found that eggplants grafted onto S. integrifolium x S. melongena rootstocks grew better

at lower temperatures (18°C to 21°C) than non-grafted plants, and Midmore et al. (1997)

observed tomato/eggplant interspecific grafts produce acceptable yields during the

Taiwan rainy season. Black et al. (2003) recommends using eggplant rootstock for

tomatoes when flooding or waterlogged soils are expected, and to select lines that are

resistant to bacterial wilt and other soil borne diseases.

25

An especially promising eggplant rootstock for tomato interspecific grafting is

Solanum torvum, or wild eggplant. S. torvum is native to the western tropics and India,

and already exhibits tolerance to the climatic pressures of tropical regions (Gousset et

al., 2005). This makes S. torvum an ideal candidate for tomato interspecific grafting in

equatorial regions, where environmental conditions can make tomato production difficult

(Max et al., 2009). There is also an established history of S. torvum for use as an

intraspecific grafting rootstock in S. melongena cultivation for its resistance to a wide

range of soil borne pathogens, including Verticillium dahlia, Ralstonia solanacearum,

Fusarium oxysporum and Meloidogyne spp. root-knot nematodes (Bletsos et al., 2003;

Gisbert et al., 2011; Singh and Gopalakrishnan, 1997). The compatibility of S. torvum as

a tomato interspecific grafting rootstock, however, has yet to be quantified. If tomato/S.

torvum compatibility were as high as commercially viable ISG rootstocks, it would allow

farmers and researchers to explore the use of tomato/S. torvum interspecific grafting for

production in areas with high risk of flood and drought stress. Thus, S. torvum was

selected as the rootstock of interest in this compatibility study, and will be tested against

other rootstocks of known high compatibility.

Uniform production of S. torvum rootstock seedlings can be challenging as a

result of low germination rate leading to poor seedling emergence and slow early growth

(Liu and Zhou, 2009). This potential hurdle may be overcome by rooting vegetative

cuttings of uniform size for use as rootstock. If there is no difference between the

compatibility of seed-sown S. torvum and vegetative propagated S. torvum in tomato

interspecific grafting, then the difficulties of seed production can be eliminated by

maintaining S. torvum stock plants for taking rootstock cuttings. Thus, the overall

objectives of this study were to assess the compatibility of S. torvum as a rootstock for

26

tomato interspecific grafting, and to determine any difference in graft compatibility based

on method of rootstock propagation.

Materials and Methods

The experiment was conducted at the University of Minnesota in Saint Paul, MN,

44.94 N and 93.09 W. In early May of 2013, 48 seeds of S.torvum were planted into a

plastic seed tray containing the soilless media ‘Sunshine Mix #8’ LC8’ (Sun Gro

Horticulture). All seed were covered with coarse vermiculite, lightly watered and placed

into a greenhouse. Greenhouse conditions were maintained at 21 C and 175 µmol PAR

light from 0700 to 1800 hours. S. torvum seeds were acquired from the Virgin Islands

Sustainable Farming Institute in St Croix, US Virgin Islands. Seeds were watered daily

until time of grafting.

Seven days after planting S. torvum seeds, 10 cuttings were taken from a stock

S. torvum plant and rooted in 4” tall pots containing the soilless media ‘Sunshine Mix #8’

LC8 (Sun Gro Horticulture). The bottom 6” of cuttings were dusted with Hormodin® 1

root inducing powder before placement into pots. Cuttings rooted in a mist house until

grafting procedures began. All leaves except meristems were removed from cuttings.

Twenty days after the planting of S. torvum seeds, all remaining seeds for the

experiment were planted using the methods stated above. This included 48 ‘Maxifort’

rootstock tomato seeds, 72 ‘CLN 3212A’ tomato seeds, and 72 ‘Celebrity’ tomato seeds.

‘Maxifort’ and ‘Celebrity’ seeds were acquired from Johnny’s Selected Seeds (955

Benton Avenue, Winslow, ME 04901), and ‘CLN3212A’ seeds were acquired from the

Asian Vegetable Research and Development Center (Shanhua, Tainan 74199, Taiwan).

27

By late July 2013, all seeds had germinated and seedlings had grown to

appropriate grafting size, the 4-5 true leaf stage (McAvoy, 2005). All S. torvum cuttings

had rooted. Cleft grafting was used for all plants; the most commonly used method for

solanaceous crops (Lee and Oda, 2003). With a razor blade, rootstocks were cut below

the cotyledon and a longitudinal cut was made 1.5 cm deep, about 75% the depth of the

stem. Scions were pruned to 1-3 leaves and the lower stem was cut into a tapered

wedge to place inside the depth cut of the rootstock (Lee and Oda, 2003). After

insertion, graft unions were wrapped with plastic parafilm to improve stability, reduce

chance of infection and ensure vascular contact (Toogood, 1999). The scion and

rootstock combinations that were joined to create the 10 different experimental

genotypes are shown in Table 1. Newly grafted plants were immediately brought into a

low light chamber with high relative humidity and a minimum of 18 degrees C at all times

(Lee and Oda, 2003). The chamber was constructed by wrapping clear and black plastic

around a PVC skeleton and placed into the greenhouse. Humidity was maintained by

sub-irrigating grafted plants on .35” deep Sure To Grow® capillary mats, which were

flushed to saturation with water every day (Gutierrez, 2008).

Each plant was evaluated daily to determine time until graft fusion and survival in

the chamber. Due to inconsistent rooting, only 8 S. torvum cuttings were available for

grafting. This resulted in “3212xS.torvum Veg” and “CelebrityxS.torvum Veg” having only

4 replications each. Evaluation involved observing changes in turgor pressure of each

plant. When scion turgor pressure was restored in the chamber, the plant was moved

outside the chamber. If the plant maintained turgor outside the chamber for 24 hours,

graft fusion was considered completed.

28

Analysis of variance (ANOVA) was used to compare the differences in survival

and days to fusion (DTF) between the 10 plants of Celebrity and 3212 genotypes.

ANOVA has been used to compare differences in graft compatibility through DTF in

perennial and annual crops (Estrada-Luna et al., 2002; Gisbert et al., 2011).

Results

A summary of average DTF and survival values for each scion genotype are

shown in Tables 2 and 3. In all Celebrity scions, the S. torvum rootstock had the highest

average DTF (12.3 days), but was not significantly higher than any other grafted

genotype. All grafted genotypes had a significantly higher DTF than the non-grafted

Celebrity control, which intrinsically had a DTF of 0 days. The Celebrity scion had the

lowest survival percentage when grafted onto rooted vegetative S. torvum rootstock

(0.5). This percentage was significantly lower than all other rootstock genotypes with the

exception of the self-grafted Celebrity genotype.

In 3212 scions, there was also no significant difference in DTF between all

grafted genotypes. The non-grafted 3212 control had a significantly shorter DTF (0 days)

than all other rootstock genotypes. Though the 3212 scion also had the lowest survival

percentage when grafted onto rooted vegetative cuttings of S. torvum (0.5), the

difference was not enough to confer statistical significance from any other rootstock

genotype.

Discussion

When comparing days to graft fusion among all rootstock genotypes, there is no

significant difference in the amount of time it takes for any successfully grafted plant to

form a healed graft union (Tables 2 and 3). S. torvum rootstock had the largest average

days to fusion for both Celebrity and 3212 scions, but the difference from other rootstock

29

genotypes was not significant . The largest average difference in days to fusion in all

grafts was between 3212xMaxifort and 3212xS.torvum Veg (2.1 days), though again this

was not statistically different from any other genotype comparison.

In both Celebrity and 3212 scions, vegetative S. torvum rootstock had a lower

survival rate to seed sown S. torvum rootstock, Maxifort, self grafted and non-grafted

rootstocks. The reason for vegetative S. torvum rootstock being less compatible than

seed sown S. torvum rootstock in interspecific grafting may be twofold. Initial

adventitious roots formed from cuttings are more adept at oxygen gas exchange and

less adept at water uptake than primary root systems (Jackson, 1955). Thus it is

possible that the scions grafted onto vegetative S. torvum lost turgor pressure and wilted

because of this diminished hydraulic capability. Also, since rootstock derived from S.

torvum cuttings would be older growth overall (with possible secondary growth), the

formation of a vascular cambium in the vegetative rootstock may have inhibited proper

graft fusion. More research is needed to determine the exact cause of reduced

compatibility in vegetative S. torvum rootstock.

Because of the low survival percentage of vegetative S. torvum rootstock with

both Celebrity and 3212 tomatoes, we do not recommended these combinations for

commercial production. Seed sown S. torvum may still be a viable option for interspecific

grafting based on the results in this experiment. If seed sown S. torvum is shown to be

compatible to a wide variety of tomato scions and an effective means of providing strong

environmental tolerances to the plant, it could be a valuable tool for growers worldwide.

This would be especially true in regions with minimal agricultural infrastructure, such as

tropical regions, where access to greenhouses and other environmentally controlled

enclosures is declining or unavailable (Schwarz et al., 2010; McElroy et al., 1990;

Sands, 1974).

30

Previous research documenting the effectiveness of seed sown S. torvum in

intraspecific grafting is promising. S. torvum rootstock confers resistance to a wide array

of environmental pressures (Bletsos et al., 2003; Gisbert et al., 2011; Singh and

Gopalakrishnan, 1997). Because of the lack of environmental tolerances in tomato

rootstock germplasm that S. torvum could provide, further exploration of S. torvum as a

flood and drought resistant rootstock for tomato scions is merited. The highly variable

germination rate of S. torvum seeds, however, would make production scheduling

difficult (Liu and Zhou, 2009). This potential hurdle may be overcome by grafting within

native distributions of S. torvum because of the vast wild population and cheap, easy

access to seed year-round.

Seed-sown S. torvum is a compatible rootstock with the two tomato scion

cultivars tested, Celebrity and CLN 3212A. Vegetative S. torvum rootstock showed

moderate compatibility as an interspecific grafting rootstock, but had a significantly

reduced grafting success rate when compared to seed sown S. torvum, Maxifort and

self-grafted rootstocks. All successfully grafted plants had similar days until graft fusion.

We recommend that the effectiveness of S. torvum rootstock in providing flood and

drought tolerances to tomato scions be explored further.

31

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AVRDC. 1990. Vegetable Production Training Manual. Asian Vegetable Research and Training Center, Shanhua, Taiwan.

Bhatt, R., N. Srinivasa Rao, and A. Sadashiva. 2002. Rootstock as a source of drought tolerance in tomato (Lycopersicon esculentum Mill.). Indian journal of plant physiology 7(4):338-342.

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Bletsos, F., C. Thanassoulopoulos, and D. Roupakias. 2003. Effect of grafting on growth, yield, and Verticillium wilt of eggplant. HortScience 38(2):183-186.

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Fernández-García, N., A. Cerdá, and M. Carvajal. 2003. Grafting, a useful technique for improving salinity tolerance of tomato?.

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Gutierrez, A. 2008. Microgreens Production with Sure To Grow(r) Pads. Sure To Grow (r), University of Vermont.

Jackson, W.T. 1955. The role of adventitious roots in recovery of shoots following flooding of the original root systems. Am. J. Bot. :816-819.

King, S.R., A.R. Davis, W. Liu, and A. Levi. 2008. Grafting for disease resistance. HortScience 43(6):1673-1676.

Lee, J.M. and M. Oda. 2003. Grafting of herbaceous vegetable and ornamental crops. Horticultural reviews :61-124.

Liu, N., X. B. Zhou, B. Zhao, Y. Lu, Li, and J. Hao. 2009. Grafting Eggplant onto Tomato Rootstock to Suppress Verticillium dahliae Infection: The Effect of Root Exudates. HortScience 44:2058-2062.

Max, J.F.J., W.J. Horst, U.N. Mutwiwa, and H. Tantau. 2009. Effects of greenhouse cooling method on growth, fruit yield and quality of tomato (Solanum lycopersicum L.) in a tropical climate. Scientia Horticulturae 122(2):179-186.

McAvoy, R. 2005. Grafting Techniques for Greenhouse Tomatoes, Nov 10, 2005.

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Singh, P. and T. Gopalakrishnan. 1997. Grafting for wilt resistance and productivity in brinjal (Solanum melongena L.). Horticultural Journal 10(2):57-64.

Toogood, A. 1999. Plant Propagation. DK Publishing, Inc, London.

Venema, J.H., B.E. Dijk, J.M. Bax, P.R. van Hasselt, and J.T.M. Elzenga. 2008. Grafting tomato ( Solanum lycopersicum) onto the rootstock of a high-altitude accession of Solanum habrochaites improves suboptimal-temperature tolerance. Environ. Exp. Bot. 63(1):359-367.

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34

Table 1. Scion and rootstock combinations analyzed for grafting compatibility. Scion Rootstock Final 'Genotype' Number of Plants

CLN 3212A none (non-grafted) 3212 10

CLN 3212A CLN 3212A 3212x3212 10

CLN 3212A ‘Maxifort®’ 3212xMaxifort 10

CLN 3212A Solanum torvum seed 3212xS.torvum 10

CLN 3212A Solanum torvum cutting 3212xS.torvum Veg 4

Celebrity none (non-grafted) Celebrity 10

Celebrity ‘Celebrity’ CelebrityxCelebrity 10

Celebrity ‘Maxifort®’ CelebrityxMaxifort 10

Celebrity Solanum torvum seed CelebrityxS.torvum 10

Celebrity Solanum torvum cutting

CelebrityxS.torvum Veg

4

Table 2. Average days to fusion and survival (%) among all ‘Celebrity’ rootstock genotypes. Letters denote statistical differences (p<0.05) within rows.

Celebrity Scion

Rootstock

Non-grafted Celebrity Maxifort S. torvum S. torvum Veg

(n) 10 10 10 10 4 Days to Fusion 0 a 12.14 b 10.5 b 12.3 b 12 b Survival % 100 a 70 ab 100 a 100 a 50 b

Table 3. Average days to fusion and survival (%) among all 3212 rootstock genotypes. Letters denote statistical differences (p<0.05) within rows.

3212 Scion

Rootstock

Non-grafted 3212 Maxifort S. torvum S. torvum Veg

(n) 10 10 10 10 4 Days to Fusion 0 a 11 b 9.9 b 11.2 b 12 b Survival % 100 80 100 80 50

35

CHAPTER 3

UTILIZATION OF INTERSPECIFIC GRAFTING FOR INCREASED FLOOD AND

DROUGHT RESISTANCE IN TOMATOES


Andrew Petran

36

Summary

Two tomato scions (‘Celebrity’ and ‘CLN3212A’) were grafted onto eggplant rootstock to

determine the effect of interspecific grafting on flood and drought tolerance of tomatoes.

Wild eggplant Solanum torvum was selected as the interspecific rootstock of interest,

and was compared against ‘Maxifort’, self-grafted and non-grafted control rootstock in

flood stress, drought stress, and optimal soil moisture conditions. Plant height, internode

length, and stomatal resistance of all scion/rootstock combinations in each

environmental condition were measured for 25 days. No significant differences in plant

height, internode length and stomatal resistance among related scion genotypes

occured in optimal conditions. In flood conditions, CelebrityxS.torvum had significantly

shorter height and internode length, and reduced visible symptoms of deoxygenation

stress. Drought conditions revealed that plants grafted on all rootstock genotypes except

S. torvum had permanently wilted by day 22, while no plants grafted onto S. torvum

wilted for the duration of the experiment. Further research is needed to determine if the

observed resistance to flood and drought conditions conferred by S. torvum would also

effect flower bud initiation, fruit set and yield.

Introduction

Improving the environmental and climatic stress tolerance of vegetable crops

through grafting is a novel approach that has been used extensively in East Asia during

the 20th century (AVRDC, 1990). In this context, grafting involves joining together two

living plant parts- a rootstock and a scion- to produce a single, living plant. Grafting is an

ideal technique for vegetable production because scions with desirable fruit-producing

traits that are also susceptible to soil-borne disease or climatic pressures can be grafted

onto rootstock that is more resistant to these pressures. The resulting union often results

37

in a more productive plant (Cohen et al., 2002; Miller et al., 2005). This solution can be

preferable to breeding resistant varieties of desirable plants because it is far less time

consuming, expensive, and technically demanding (Sleper et al, 1991). Since grafting

enhances environmental tolerances without altering the environment itself, it would

intrinsically reduce the input requirement of pesticides for optimal production. Thus, this

technique would not tax the minimal agricultural infrastructure that exists in many tropical

regions today (EarthTrends, 2003).

Grafting vegetables onto tolerant rootstock of its own species is the most

common grafting technique since intraspecific compatibility is high (Black et al., 2003;

Palada and Wu, 2008; Rivard and Louws, 2008). While intraspecific grafting has been

proven to increase resistance to various environmental pressures such as flood,

drought, cold, heat and pathogen stress, in some cases the transferred tolerance is not

strong enough, or a certain desired environmental tolerance does not yet exist within the

rootstock germplasm of that species (Bloom et al., 2004; Sanders and Markhart, 1992;

Venema et al., 2008; Zijlstra and Nijs, 1987). However, vegetable species with certain

environmental susceptibilities will sometimes have grafting-compatible relatives within

the same family that possess a natural resistance to that stress. Thus, interspecific

grafting can be used to broaden rootstock diversity when environmental pressures

surpass the advantages that can be provided by intraspecific grafting alone.

The utilization of interspecific grafting as an alternative, low-input means of vegetable

production in regions with high environmental pressures is promising, and merits further

exploration. Interspecific grafting has been used successfully for many vegetables in the

curcubit and solanaceous families (King et al., 2010). After grafting Solanum

melongena L. eggplant scions onto a verticillium wilt resistant tomato rootstock Lydl, Liu

and Zhou (2009) found a 0% incidence of the disease on grafted eggplant, compared to

38

a 68.3% incidence on non-grafted controls. Allelopathic chemicals were also found in the

tomato rootstock exudates, inhibiting spore germination and mycelium growth. Davis et

al. (2008) states that watermelon grafted onto the rootstock of bottle gourd (Lagenaria

siceraria) confer significant resistance to Fusarium spp.

The identification of an ideal rootstock for interspecific grafting compatibility trials

can be achieved by surveying available research for a history of compatibility, and giving

preference to plants native to the region the grafted crop will be cultivated. This way, the

likelihood of compatible rootstock having appropriate environmental tolerances to be

transferred to the non-native scion may be increased. If a compatible native rootstock

also with a history of successful use can be found, then probability of successful trials

grows even more.

Tomatoes are one of the most lucrative cash crops worldwide, but they are

especially sensitive to excessive flooding or drought, making them difficult to produce in

tropical regions (EarthTrends, 2003; Max et al., 2009). Multiple accessions of tomato

rootstock are in use commercially to confer temperature and salinity tolerances, but to

date no tomato rootstock has significant resistance to flood conditions (Bhatt et al., 2002;

Fernandez-Garcia et al., 2003; Schwarz et al., 2010). Thus, the identification of a flood

tolerant interspecific graft would help fill the gap of flood tolerance in tomato production.

One potential candidate is eggplant (Solanum melongena), a highly resilient and closely

related solanaceous plant to tomato with many cultivars whose roots can survive for

several days underwater.

There is a history of tomato/eggplant interspecific grafting to mitigate unfavorable

climatic conditions, and vice versa. Okimura et al. (1986) found that eggplants grafted

onto S. integrifolium x S. melongena rootstocks grew better at lower temperatures (18°C

39

to 21°C) than non-grafted plants. Midmore et al. (1997) observed tomato/eggplant

Interspecific grafting produces acceptable yields during the rainy season. Black et al.

(2003) recommends using eggplant rootstock for tomatoes when flooding or waterlogged

soils are expected, and to select lines that are resistant to bacterial wilt and other soil

borne diseases. The AVRDC has found that eggplant accessions EG195 and EG203 are

compatible with most tomato scions and resistant to flooding, bacterial wilt, fusarium wilt

and root-knot nematode, and Chetelat and Peterson (2003) have identified tomato

accession ‘Hawaii 7998’ as broadly compatible with distantly related solanaceous crops,

such as eggplant and pepper. No tomato/eggplant interspecific grafting trials have been

performed analyzing for drought tolerance, but Pena and Hughes state that some

eggplant genotypes are drought tolerant and eggplant rootstocks may therefore provide

protection against soil moisture stress (2007). This is likely due to eggplant being more

effective at water uptake than tomato root systems (Schwarz et al., 2010).

A variety of wild eggplant, Solanum torvum, has been selected as a rootstock

candidate for tomato/eggplant interspecific grafting in this experiment. Solanum torvum

is native to the western tropics and India, and already exhibits tolerance to the climatic

pressures of tropical regions (Gousset et al., 2005). There is also an established history

of S. torvum for use as a rootstock in S. melongena cultivation for its resistance to a

wide range of soil borne pathogens, including Verticillium dahlia, Ralstonia

solanacearum, Fusarium oxysporum and Meloidogyne spp. root-knot nematodes

(Bletsos et al., 2003; Gisbert et al., 2011; Singh and Gopalakrishnan, 1997). Uniform

production of S. torvum rootstock seedlings can be challenging as a result of low

germination rate leading to poor seedling emergence and slow early growth (Liu and

Zhou, 2009). This potential hurdle may be overcome by rooting vegetative cuttings of

stock plants for use as rootstock, or by grafting within native distributions of S. torvum,

40

such as the U.S. Virgin Islands, because of the vast wild population and cheap, easy

access to seed year-round.

If tomato crops grafted onto S. torvum produces a plant able to withstand flood

and drought, then it could be utilized as a low-input tool to optimize production in tropical

regions such as the U.S. Virgin Islands. Possible advantages of growing rootstocks in

these areas include diversifying local cultivar availability and low-input season extension.

Extending the season for local produce allows for off-season price premiums, sometimes

as high as 50% (Jett, 2006; Rowley, 2010; Rowley et al., 2010).

Stomatal conductance and resistance is the quantitative measurement of plant

gas exchange on the leaf. Because water leaves the stomata during this gas exchange,

stomatal conductance and resistance has also been used to measure flood tolerance,

drought tolerance and overall water use efficiency in plants (Kato et al., 2001; Sivritepe

et al., 2005). A porometer measuring stomatal resistance can be used to determine the

physiologic response of each genotype under controlled moisture conditions. Porometer

readings are taken on newly unfurled leaves at the same time of day, as changes in age

of the leaf and time of day can alter the consistency of resistance readings and confound

statistical analysis (Ferreira and Katerji, 1992).

The tomato rootstock ‘Maxifort’ is a commercial standard for grafting tomato

(Rivard and Louws, 2008), and was utilized as a control rootstock. Non-grafted Celebrity

and 3212 were also used as a control. Two tomato scions, ‘Celebrity’ and ‘CLN 3212A’

(3212) were chosen for this project. Celebrity was chosen because of its wide use, and

3212 was chosen because the Asian Vegetable Research and Development Center’s

classification as a heat tolerant cultivar. If interspecific grafting onto S. torvum rootstock

would confer moisture tolerance to either cultivar, this technique could be used to

41

increase the adaptation of this vegetable crop. The objective of this research was to

determine whether S. torvum as an interspecific rootstock can impart flood and drought

tolerance of tomato scions.



44.94 N and 93.09 W. In June of 2012, 48 seeds of S.torvum were planted into a 48-

count plastic seed tray containing the soilless media ‘Sunshine Mix #8’ LC8’ (Sun Gro


into a greenhouse maintained at 21°C and 175 µmol PAR light from 0700 to 1800 hours.

S. torvum seeds were acquired from the Virgin Islands Sustainable Farming Institute in

St Croix, US Virgin Islands. Seeds were watered daily until time of grafting.

Twenty days after the planting of S. torvum, all remaining seeds for the

experiment were planted using the methods stated above. This included 48 ‘Maxifort’

rootstock tomato seeds, 72 ‘CLN 3212A’ tomato seeds, and 72 ‘Celebrity’ tomato seeds.

‘Maxifort’ and ‘Celebrity’ seeds were acquired from Johnny’s Selected Seeds (955

Benton Avenue, Winslow, ME 04901), and ‘CLN3212A’ seeds were acquired from the

Asian Vegetable Research and Development Center (Shanhua, Tainan 74199, Taiwan).

In early July 2012, all seeds had germinated and seedlings had grown to

appropriate grafting size, the 4-5 true leaf stage (McAvoy, 2005). Plants were grafted

using the cleft grafting technique, which is most commonly used on solanaceous crops

(Lee and Oda, 2003). With a razor blade, rootstocks were cut below the cotyledon and a

longitudinal cut was made 1.5 cm deep, about 75% the depth of the stem. Scions were

pruned to have 1-3 leaves and the lower stems were cut into a tapered wedge to place

42

inside the depth cut of the rootstock (Lee and Oda, 2003). After insertion, graft unions

were wrapped with plastic parafilm to improve stability, reduce chance of infection and

ensure vascular contact (Toogood, 1999). The scion and rootstock combinations that

were joined to create the 8 different experimental genotypes are shown in Table 1.

Newly grafted plants were immediately brought into a low light environment with high

relative humidity and a minimum of 18 degrees C at all times (Lee and Oda, 2003). This

chamber was constructed by wrapping clear and black plastic around a PVC skeleton

and placed into the greenhouse. Humidity was maintained by sub-irrigating grafted

plants on .35” deep Sure To Grow® capillary mats, which were flushed to saturation with

water every day (Gutierrez, 2008). After approximately seven days the plants were

removed from the chamber to be grown on in experimental conditions.

After graft unions had healed, plants were taken out of the healing chamber and

potted into 6” pots filled with LC8 media (Sun Gro Horticulture). Plants were divided into

3 different moisture treatments (Drought Stress, Flood Stress and Optimal Conditions),

each treatment having 3 replications of the 8 different combinations (3212, 3212x3212,

3212xS.torvum, 3212xMaxifort, Celebrity, CelebrityxCelebrity, CelebrityxS.torvum,

CelebrityxMaxifort). Plants were arranged in a randomized complete block design, with

each block representing a different moisture treatment.

Optimal soil moisture conditions were maintained by calculating how many mL of

water were needed to increase soil moisture in the pot by 1%. This was done by, first,

measuring the mL of water lost from a 6” pot with LC8 soilless media 24 hours after

saturation, and dividing it by the percentage of soil moisture lost over the same time

period. With this figure, each plant could be measured for soil moisture every day and be

given the exact amount of water needed to bring the soil moisture content back to the

optimal level (known as container capacity) observed 24 hours after saturation (White

43

and Mastalerz, 1966). It was calculated that container capacity for the media in 6” pots

was 34% soil moisture. Soil moisture was read daily with an SM100 Soil Moisture

Sensor plugged into a Watchdog 1000 Series Micro Station.

Drought stress conditions were maintained by using the same daily watering

technique used for optimal conditions, except water was given to maintain soil moisture

in between 1% and 2%, unlike the optimal soil moisture treatment, which in this

experiment was soil moisture of 34%.

Flooded conditions were created by placing the plants directly into plastic basins

half the height of the pots, and filling the basins with water every day. This ensured that

the soil moisture levels were at saturation for the duration of the experiment.

Immediately after potting and establishment of each environmental condition,

plants were measured for morphological and physiological changes over a 25-day

period. Plant height (cm), number of nodes, internode length and stomatal resistance

(mmol/m2/s) readings were taken on day 1, 5, 10, 15, 20, and 25, along with plant

survival. Stomatal resistance was measured with a Delta T® AP4 porometer.

Resistance readings were taken on the terminal leaflet of the youngest fully-unfurled

branch each measurement day at 1 pm.

On the final day, fresh weight, dry weight and leaf area (cm2) measurements

were taken. Leaf area was measured using a Li-Cor LI-3100 area meter prior to placing

plants in a Hot Pack® drying oven at 170 degrees F for 48 hours. Dry weight was taken

after XXX days.

Data was subjected to multiple methods of statistical analysis. Analysis of

variance (ANOVA) was used to compare the differences in stomatal resistance between

the four Celebrity and 3212 genotypes in each environmental condition (flood, drought,

44

optimal) on each measurement day. ANOVA has been used to compare differences in

stomatal conductance in tomatoes exposed to different categorical treatments (Sivritepe

et al., 2005). ANOVA was also used to compare the differences in final plant height and

internode lengths (day 25) between the four Celebrity and 3212 genotypes in each

environmental condition. Linear regression analysis was used to determine the

significance of correlation between dry weight and leaf area in each environmental

condition, and also to determine the significance of a stomatal resistance regression of

the four Celebrity and 3212 genotypes in each environmental condition (flood, drought,

optimal) over time. Rootstock genotype may have an effect on the resistance of the

scion in stressed conditions (Borel et al., 2001). Statistical significance for ANOVA and

regression was calculated at the p < 0.05 level. All analyses were carried out using the

statistical program R.

Results

ANOVA- Plant Height and Internode Length

Under optimal conditions, the final plant height of 3212xS.torvum was

significantly different (p < 0.05) than 3212x3212, and CelebrityxS.torvum was not

significantly different than any other Celebrity genotype (Figure 1). There was no

significant difference in internode length among any genotypes in optimal conditions

(Figure 2). In flood stress conditions, final plant height of 3212xS.torvum was not

significantly different than any other 3212 genotypes, and CelebrityxS.torvum was

significantly different than all other Celebrity genotypes (Figure 1). Final internode length

of CelebrityxS.torvum was significantly different than Celebrity and CelebrityxCelebrity

45

(Figure 2). In drought stress conditions, no genotype had a significantly different plant

height or internode length than any other genotype within its group (Figures 1 and 2).

When analyzing stomatal resistance, data taken from day 20 was used as all

plants in the drought treatment were dead by day 25. In optimal conditions,

3212xS.torvum was significantly different than 3212xMaxifort (Figure 3). In both flood

and drought stress conditions there was no significant difference between any related

genotypes (Figure 3).

Linear Regression Analysis

A highly significant correlation between dry weight and leaf area among all 3

treatments was determined with regression analysis. Correlation in optimal conditions

had a p-value of 0.047, compared to a correlation in drought conditions of p=0.046 and

flooded conditions of p=0.087. When all treatments were combined, correlation between

dry weight and leaf area had an R-squared value of .71 with a p-value of 2.2e-16 (Figure

4). No significant differences among the average stomatal resistance of any related

genotypes over time occurred, but certain observable trends could be seen. In optimal

conditions there was general inconsistency regarding which genotypes had the highest

and lowest graft combinations as time progressed, but in flooded conditions

3212xS.torvum consistently had the highest stomatal resistance and in drought stress

CelebrityxS.torvum consistently had the lowest stomatal resistance.

Discussion

Optimal Conditions

Under optimal conditions both final plant height and internode length of 3212 and

Celebrity genotypes are reduced when grafted onto S.torvum rootstock, though the only

46

statistically significant difference was between 3212xS.torvum and 3212x3212. This

observable reduction in height may be due to the grafting procedure itself. Time from

sowing to transplant of grafted tomatoes is 30 to 33 days, while non-grafted tomatoes

take less time, 14 to 21 days (Black et al., 2003; McAvoy and Giacomelli, 1985). In the

procedure of this experiment, scions and non-grafted controls were sown on the same

day so non-grafted plants would have a height advantage over grafted transplants.

Intraspecific grafted plants were able to make up for this deficit by day 25, but 3212xS.

torvum and CelebrityxS. torvum still had the lowest mean plant height and internode

length, most likely due to the increased time it takes for graft unions to fuse in S. torvum

interspecific grafting (Masayuki et al., 2005). In the future, if scion sowing for S. torvum

interspecific grafting is started before other genotypes, it would likely offset this initial

height difference. Also, since internode length of all related scion genotypes were so

similar, it is unlikely that this reduction in height was due to environmental stress (Figure

2).

No Celebrity genotype had a significantly different stomatal resistance than other

Celebrity genotypes over the course of the experiment, but in day 20 3212xS. torvum

had a significantly higher resistance than 3212xMaxifort. This may have led to

3212xMaxifort having a higher median plant height than 3212xS. torvum by the end of

the experiment (Figure 1), but by day 25 there was no difference in stomatal resistances.

The overall lack of significant differences imply that in optimal moisture

conditions, grafting onto S. torvum or Maxifort rootstock does not confer a distinct

morphological or physiological growth advantage or disadvantage compared to each

other or to an non1grafted control. These results differ from those found by Fernandez-

Garcia et al. (2002) that grafted tomatoes experience superior stomatal conductance

compared to non-grafted controls of the same cultivar, even in optimal conditions. Since

47

this project did not quantify flowering, total or marketable yield, we cannot determine

whether this insignificant difference in growth also results in yield differences among

genotypes. A second trial of this experiment, conducted in February 2013, yielded the

same optimal condition results among genotypes as the initial 2012 trial.

Photograph 1 shows the observable differences within each genotype. Previous

research has found that even without the presence of temperature, moisture or pathogen

stress grafted tomatoes can still lead to increased yields compared to non-grafted

controls, especially in older heirloom cultivars (Rivard and Louws, 2008). Thus, grafting

tomatoes may be a financially viable cultural practice even in optimal production

conditions. Possible explanations for vegetable yield increase in optimal conditions

include increased water and macronutrient uptake by vigorous rootstock genotypes

(Ruiz and Romero, 1999; Yetisir and Sari, 2003). Fernandez-Garcia et al. (2002) found

that certain rootstocks can improve the stomatal conductance of tomato scions.

Fernandez-Garcia et al. (2002) found that certain rootstocks can improve the stomatal

conductance of tomato scions and Leonardi and Giuffrida (2006) observed increased

phosphorus and calcium uptake with particular rootstock genotypes as well.

Flood Conditions

Flood stress causes various physiological and morphological responses in

tomatoes. Waterlogged soils inhibit oxygen and nutrient uptake, which causes leaf

chlorosis and necrosis, starting in mature leaves and slowly making it’s way up the plant

(Ezin et al., 2010). Waterlogged soils sometimes cause the underside of tomato leaves

to turn purple as a result of phosphorus deficiency (Dumas, 1989). Flood stressed

tomatoes also temporarily close their stomata, until the formation of adventitious roots as

a mechanism to alleviate the stress (Aloni and Rosenshtein, 1982; Kozlowski, 1984).

48

The formation of adventitious roots increase oxygen uptake, and stomatal conductance

levels return to near normal. In this project, adventitious roots were observed in all

genotypes, along the soil line and in some cases on the scion at the point of graft union.

ANOVA testing revealed that although grafting didn’t appear to have an effect on 3212

genotypes in flooded conditions, grafting Celebrity onto S. torvum had a profound effect

when soils were waterlogged. CelebrityX S. torvum had a significantly shorter plant

height and internode length than all other Celebrity genotypes (Figures 1 and 2). Also,

when comparing treatment differences of specific genotypes, almost all genotypes

experience increased or similar plant heights and internode lengths when flood stress is

applied compared to optimal conditions, while CelebrityXS. torvum decreases in these

categories. Thus, flood stress resulted in taller or leggier plants in most tomato

genotypes, but result in a shorter, more compact plant when grafted onto S. torvum.

Visible differences in leaf color can be observed as well (Photograph 2). As

mentioned before, flood stress symptoms in tomatoes include chlorosis and necrosis of

mature leaves along with purple undersides of leaves and veins (Ezin et al., 2010). All

these symptoms were seen to a high degree in every genotype with the exception of

CelebrityxS. torvum, where mature leaves were still green and had limited purpling of

veins. Visual differences show that while waterlogged soils lead CelebrityxS. torvum to

be shorter and more compact, fewer visual symptoms were noted when compared to all

other genotypes. Further exploration should include a leaf tissue analysis of all

genotypes, so more precise reasons for visual differences can be quantified. A second

trial of this experiment, conducted in February 2013, yielded the same flood tolerance

results as the initial 2012 trial.

Drought Conditions

49

There was no statistical difference among related scion genotypes in any of the

measurements taken in drought conditions. As in other treatments, CelebrityxS. torvum

had the lowest median plant height and internode length, but unlike in optimal or flooded

conditions 3212xS. torvum had the highest median plant height and internode length

under drought stress (Figures 1 and 2). This trend difference may be attributed to 3212

being known as a heat tolerant tomato variety by its distributer, the Asian Vegetable

Research and Development Center. It has been shown before that increasing heat

tolerance in tomato can result in an increase in water use efficiency (Lukic et al., 2012).

Stomatal resistance of all genotypes in drought stress is high when compared to

optimal and flooded conditions, although there is no significant difference among related

genotypes within the drought treatment (Figure 3). These results are consistent with the

literature, that stomatal resistance is known to increase when tomatoes are exposed to

drought stress conditions to conserve water (Camejo et al., 2005; Sobeih et al., 2004).

Photograph 3 shows the differences among scions grafted onto S. torvum and

other rootstocks in drought stress. 3212 scions grafted onto S torvum have noticeably

more turgor pressure than 3212, Maxifort and non-grafted rootstock. In Celebrity, all

rootstock except S. torvum reached permanent wilting point by day 22. Celebrity scions

reaching permanent wilting before 3212 may be attributed to the increased heat

tolerance in 3212 raising water use efficiency as well (Lukic et al., 2012). A second trial

of this experiment, conducted in February 2013, yielded the same drought tolerance

results as the initial 2012 trial.

A thorough review of the literature yielded no available research exploring

rootstock induced drought tolerance in tomatoes, so the exact mechanism leading to the

tolerance observed in this experiment is not yet known. In apples, dwarfing and semi-

50

dwarfing rootstock use less water due to higher leaf-specific, soil-stem hydraulic

resistance (Cohen and Naor, 2002). Since S. torvum appears to have a dwarfing effect

on both Celebrity and 3212, it is possible the same phenomenon may be occurring here.

In this experiment, we found that S. torvum rootstock does not effect plant height,

internode length or stomatal resistance of tomato scions Celebrity and CLN 3212A in

optimal moisture conditions. In flood conditions, CelebrityxS.torvum had significantly

shorter height and internode length, and reduced visible symptoms of deoxygenation

stress. Drought conditions revealed that plants grafted on all rootstock genotypes except

S. torvum had permanently wilted by day 22, while no plants grafted onto S. torvum

wilted for the entirety of the experiment. Based on these findings, we recommend using

S. torvum as a rootstock for interspecific tomato grafting to increase drought tolerance.

This practice would be particularly useful for producers located in regions of

considerable drought stress.

Future research could investigate root architecture of S. torvum rootstock against

rootstocks not shown to be drought tolerant, plant nutrient differences among genotypes

via plant tissue analysis, and chemical root-to-shoot signaling that controls water use

efficiency in S. torvum versus other rootstock.

51


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Sivritepe, H.Ö., N. Sivritepe, A. Eriş, and E. Turhan. 2005. The effects of NaCl pre-treatments on salt tolerance of melons grown under long-term salinity. Scientia horticulturae 106(4):568-581.

Sleper, D.A., T.C. Barker, and P.J. Bramel-Cox. 1991. Plant Breeding and Sustainable Agriculture: Considerations for objectives and methods. Crop Society of America, Madison, WI.

Sobeih, W.Y., I.C. Dodd, M.A. Bacon, D. Grierson, and W.J. Davies. 2004. Long-distance signals regulating stomatal conductance and leaf growth in tomato (Lycopersicon esculentum) plants subjected to partial root-zone drying. J. Exp. Bot. 55(407):2353-2363.


54



White, J. and J. Mastalerz. 1966. Soil moisture as related to container capacity.

Yetisir, H. and N. Sari. 2003. Effect of different rootstock on plant growth, yield and quality of watermelon. Animal Production Science 43(10):1269-1274.


55

Table 1. 8 different scion and rootstock combinations were analyzed for morphological and physiological responses to different environmental pressures.

Scion Rootstock Final 'Genotype'

# Replications per Environmental Treatment

CLN 3212A none (non-grafted) "3212"

3

CLN 3212A CLN 3212A "3212x3212" 3 CLN 3212A Maxifort® "3212xMaxifort" 3

CLN 3212A Solanum torvum "3212xS.torvum" 3

Celebrity none (non-grafted) "Celebrity"

3

Celebrity Celebrity "CelebrityxCelebrity"

3

Celebrity Maxifort® "CelebrityxMaxifort"

3

Celebrity Solanum torvum "CelebrityxS.torvum"

3

56

Figure 1. Boxplot of plant height (cm) of all genotypes in each treatment, day 25. Star

indicates a significant difference (p<0.05) from all related scion genotypes in that

condition.

*

57

Figure 2. Boxplot of internode length (cm) of all genotypes in each treatment, day 25.

Star indicates a significant difference (p<0.05) from all related scion genotypes in that

condition.

*

58

Figure 3. Boxplot of stomatal resistance (mmol/m2/s) of all genotypes in each condition,

day 25.

59

Figure 4. Scatter plot showing the correlation of dry weight (g) to leaf area (cm2) of all

treatments.

60

Photograph 1. Randomized selection of each genotype in optimal conditions, Trial 1,

Day 22.

61

Photograph 2. Randomized selection of each genotype in flooded conditions, Trial 1,

Day 22. Star placed above CelebrityxS. torvum to indicate significant difference in plant

height and internode length.

62

Photograph 3. Randomized selection of each genotype in drought conditions, Trial 1,

Day 22.

63

CHAPTER 4

UTILIZATION OF INTERSPECIFIC GRAFTING FOR INCREASED HEAT TOLERANCE IN TOMATOES


Andrew Petran

64

Summary

Two tomato scions (‘CLN 3205A’ and ‘CLN3212A’) were grafted onto eggplant

rootstock to determine the effect of interspecific grafting on heat tolerance of tomatoes.

Wild eggplant Solanum torvum was selected as the interspecific rootstock of interest,

and was compared against an non-grafted control rootstock in plant growth chambers

set at optimal (26 C day, 20 C night) and supraoptimal temperatures (37 C day, 28 C

night) for tomato vegetative growth. Plant height, internode length, and stomatal

resistance of all genotypes in each environmental condition were measured for 35 days.

Changes in plant height and internode length of tomatoes placed in supraoptimal

temperatures was similar for both S. torvum and non-grafted rootstock, but non-grafted

tomatoes experienced a significant decrease in stomatal resistance when placed in

supraoptimal temperatures, while tomatoes grafted onto S. torvum did not. Further

research is needed to determine if the observed stomatal stability in supraoptimal

temperature conditions of grafted plants could lead to an increase in yield through flower

initiation and fruit set.

Introduction

Temperature stresses, both too high and too low, have been shown to lead to

inhibited plant growth and development, wilt, necrosis, and reduced overall yield of

curcubit and solanaceous crops, including tomato (Ahn, 1999). Specifically, supra-

optimal temperatures have observed deleterious effects on crops as well, including

growth reduction, decrease in photosynthetic capacity accompanying increased

respiration, osmotic damages and inhibited ion uptake/transport (Wang et al., 2003).

Such conditions are applicable in many areas of the world, but of special concern to

growers located in the lowland tropics (Palada and Wu, 2006).

65

Improving the environmental and climatic stress tolerance of vegetable crops

through grafting is a novel approach that has been in used extensively in East Asia

during the 20th century (AVRDC, 1990). In this context, grafting involves joining together

two living plant parts- a rootstock and a scion- to produce a single, living plant. Grafting

is an ideal technique for vegetable production because scions with desirable fruit-

producing traits that are also susceptible to soil-borne disease or climatic pressures can

be grafted onto rootstock that is more resistant to these pressures. The resulting union

often results in a more productive plant (Cohen et al., 2002; Miller et al., 2005). Grafting,

thus, this technique would not tax the minimal agricultural infrastructure that exists in

many tropical regions today (EarthTrends, 2003).

Tomatoes are one of the most lucrative cash crops worldwide, but many cultivars

and genotypes are affected when exposed to supraoptimal temperatures similar to

conditions in the USVI (Abdalla and Verderk, 1968; EarthTrends, 2003). Most tomatoes

grow under optimum day/night temperatures of 25 degrees Celsius (Max et al., 2009),

however some tomato genotypes are tolerant at higher temperatures. Abdul-Baki (1991)

found that several tomato lines genetically selected for heat tolerance produced a higher

yield in high temperature conditions (38-40° Celsius) than when they were grown in

normal field conditions (26-28° Celsius). These selected genotypes would likely produce

well under high heat stress, but if a producer wanted to confer heat tolerance to other,

exotic heirloom varieties, interspecific grafting may be a viable option.

Explorations of grafted plants growing in expanded temperature conditions on

total yield have shown highly variable results. Initial trials by Okimura et al. (1986) and

Bulder et al. (1987) on Cucurbitaceae and Solanaceae showed that different scion-

rootstock combinations don’t respond with significantly different yields in suboptimal

temperatures, but subsequent trials have identified rootstocks that lead to higher overall

66

yields in tomato, cucumber and watermelon (Ahn et al., 1999; Davis et al., 2008;

Tachibana, 1982; Ziljstra and Nijs, 1987). Not surprisingly, all grafted combinations that

show sub and supra-optimal temperature tolerances are those with rootstocks with wide

optimal temperature thresholds (Schwarz et al, 2010).

In supraoptimal temperature conditions, research has shown that grafting

tomatoes onto heat-tolerant tomato rootstock increases vegetative growth, but with no

significant difference in yield compared to non-grafted controls (Abdelmageed and

Gruda, 2009). However, using eggplant as an interspecific grafting rootstock may be a

more promising candidate for supraoptimal temperature conditions, since eggplants are

more adapted to live in hot, arid climates (Abdelmageed and Gruda, 2009; Schwarz et

al., 2010). Indeed, Wang et al. (2006) observed yield increases of 10% on eggplant

when grafted onto heat-tolerant eggplant rootstock. If similar results could be observed

for tomato-eggplant interspecific grafting, it could increase local tomato production,

cultivar availability, and growing season in areas of supraoptimal temperature for tomato.

A species of wild eggplant, Solanum torvum, has been selected as a rootstock

candidate for tomato/eggplant interspecific grafting in this experiment. Solanum torvum

is native to the western tropics and India, and exhibits tolerance to the climatic pressures

of tropical regions (Gousset et al., 2005). There is also an established history of S.

torvum for use as a rootstock in S. melongena cultivation for its resistance to a wide

range of soil borne pathogens, including Verticillium dahlia, Ralstonia solanacearum,

Fusarium oxysporum and Meloidogyne spp. root-knot nematodes (Bletsos et al., 2003;

Gisbert et al., 2011; Singh and Gopalakrishnan, 1997). If evidence of increased

temperature tolerance could be confirmed as well, it would make S. torvum an even

more appealing candidate for commercial interspecific grafting production in the tropics.

Uniform production of S. torvum rootstock seedlings can be challenging due to low

67

germination rate leading to poor seedling emergence and slow early growth (Liu and

Zhou, 2009).

Stomatal conductance and resistance is the quantitative measurement of plant

gas exchange of the leaf. Because water leaves the stomata during this gas exchange,

stomatal conductance and resistance has also been used to measure flood tolerance,

drought tolerance and overall water use efficiency in plants (Kato et al., 2001; Sivritepe

et al., 2005). A porometer was used in this project to measure stomatal resistance of

each genotype under controlled temperature conditions. Porometer readings are taken

on newly unfurled leaves at the same time of day since changes in leaf age and time of

day can alter the consistency of resistance readings and confound statistical analysis

(Ferreira and Katerji, 1992).

Two tomato scions, ‘CLN 3205A’ (3205) and ‘CLN 3212A’ (3212) were chosen

for this project. These cultivars were chosen because of their classification as heat

tolerant cultivars by the Asian Vegetable Research and Development Center. If

interspecific grafting onto S. torvum rootstock would confer moisture tolerance to either

cultivar, this technique could be used to increase the adaptation of this vegetable crop.

Non-grafted Celebrity and 3212 will also be tested to serve as an unaltered, general

control for statistical comparison.

If tomato crops grafted onto S. torvum are able to confer supraoptimal

temperature tolerances that are greater than other available rootstock and non-grafted

tomatoes, then it could possibly be utilized as a low-input tool to optimize production in

regions of high temperature stress. The objective of this study, thus, was to determine

the effect of S. torvum as an interspecific grafting rootstock for improving the heat

tolerance of tomatoes.

68



44.94 N and 93.09 W. In December of 2012, 24 seeds of S.torvum were planted into a

48-count seed tray containing the soilless media ‘Sunshine Mix #8’ LC8’ (Sun Gro


into a greenhouse. Greenhouse conditions were maintained at 21° C and 175 µmol PAR

light from 0700 to 1800 hours. S. torvum seeds were acquired from the Virgin Islands

Sustainable Farming Institute in St Croix, US Virgin Islands. Seeds were watered daily

until time of grafting.

Twenty days after the planting of S. torvum, all remaining seeds for the

experiment were planted using the methods stated above. This included 24 ‘CLN 3205’

tomato seeds and 24 ‘CLN 3212A’ tomato seeds. Scion tomato seeds were acquired

from the Asian Vegetable Research and Development Center.

In early January 2013, all seeds had germinated and seedlings had grown to

appropriate grafting size, the 4-5 true leaf stage (McAvoy, 2005). Plants were grafted

using the cleft grafting technique, which is most commonly used on solanaceous crops

(Lee and Oda, 2003). With a razor blade, rootstocks were cut below the cotyledon and a

longitudinal cut was made 1.5 cm deep, about 75% the depth of the stem. Scions were

pruned to 1-3 leaves, and the lower stem was cut into a tapered wedge to place inside

the depth cut of the rootstock (Lee and Oda, 2003). After insertion, graft unions were

wrapped with plastic parafilm to improve stability, reduce chance of infection and ensure

vascular contact (Toogood, 1999). The scion and rootstock combinations that were used

in experiment are explained in Table 1. Newly grafted plants were immediately brought

into a low light environment with high relative humidity and a minimum of 18° C at all

69

times (Lee and Oda, 2003). This chamber was constructed by wrapping clear and black

plastic around a PVC skeleton and placed into the greenhouse. Humidity was

maintained by sub-irrigating grafted plants on 0.35” deep Sure To Grow® capillary mats,

which were flushed to saturation with water every day (Gutierrez, 2008). After

approximately seven days the plants were removed from the chamber to be grown on in

experimental conditions, described below.

Plants were placed in plant growth chambers and measured for morphological

and physiological changes over a 35-day period. All environmental conditions in both

growth chambers were the same, with the exception of temperature. One growth

chamber was set at 26° C day, 20° C night (designated ‘control’) and the second

chamber was set at 37° C day, 28° C night (designated ‘hot’). Plant height (cm), number

of nodes, internode length and stomatal resistance (mmol/m2/s) readings were taken on

day 1, 5, 15, 25, and 35, along with plant survival. Stomatal resistance was measured

with a Delta T® AP4 porometer. Resistance readings were taken on the terminal leaflet

of the youngest fully unfurled branch each measurement day at 1 pm, as changes in age

of the leaf and time of day can alter the consistency of resistance readings and confound

statistical analysis (Ferreira and Katerji, 1992).

On the final day, fresh weight, dry weight and leaf area (cm2) readings were

taken as well. Dry weight was taken after plants had been placed in a Hot Pack® drying

oven at 170 degrees F for 48 hours, and leaf area was measured using a Li-Cor LI-3100

area meter. All measurements for this set of replications were finished by February

2013.

Data was subjected to multiple methods of statistical analysis. Analysis of

variance (ANOVA) was used to compare the differences in stomatal resistance between

all replications of the two 3205 and 3212 genotypes in each environmental condition

70

(control, hot) on each measurement day. ANOVA has been used to compare differences

in stomatal conductance in tomatoes exposed to different categorical treatments

(Sivritepe et al., 2005). ANOVA was also used to compare the differences in final plant

height and internode lengths (day 35) between the two 3205 and 3212 genotypes in

each environmental condition. Linear regression analysis was used to determine the

significance of correlation between dry weight and leaf area in each environmental

condition, and also to determine the significance of a stomatal resistance regression of

the two 3205 and 3212 genotypes in each environmental condition over time. Rootstock

genotype may have an effect on the resistance of the scion in stressed conditions (Borel

et al., 2001). Statistical significance for ANOVA and regression was calculated at the p <

0.05 level. All analyses were carried out using the statistical program R.

Results

Control conditions with 3205 was the tallest plant on average (72 cm), and was

significantly taller than both 3212 and 3212xS.torvum, but not significantly taller than

3205xS.torvum (Table 2). There were no significant differences in average plant height

among any of the genotypes in hot conditions. 3205 and 3205xS.torvum were

significantly shorter in hot conditions than in control conditions (Table 3). There were

also no significant differences in internode length among any genotypes within each

environmental condition, but when comparing the same genotype in different conditions,

both 3205 and 3205xS.torvum had longer average internode lengths in control

conditions than in hot conditions (Table 4).

Analyzing stomatal resistance revealed that all average resistances of genotypes

were higher in control conditions than in hot conditions, but those differences were only

71

significant for non-grafted 3205 and 3212 genotypes (Table 3). There was no statistical

difference among scion genotypes in the same environmental condition.

Discussion

Regardless of temperature conditions, grafting a genotype onto S. torvum

rootstock usually reduced average plant height compared to the non-grafted control, with

the exception of the 3212 scion in control conditions. This observable yet insignificant

reduction in height may be due to the grafting procedure itself. Time from sowing to

transplant of grafted tomatoes is 30 to 33 days, while non-grafted tomatoes take less

time, 14 to 21 days (Black et al., 2003; McAvoy and Giacomelli, 1985). In the procedure

of this experiment, plants destined to be grafted as well as non-grafted control seeds

were sown on the same day, so it is understandable that non-grafted plants would have

a height advantage at the time of grafted transplant. The reason for 3212xS.torvum

having a significantly higher average plant height than 3212 in control conditions cannot

be confirmed, but it is possible that 3212 is a more compatible scion for S. torvum

interspecific grafting than 3205.

Interspecific graft compatibility is highly variable among genotypes and difficult to

predict; the degree of taxonomic affinity necessary for compatibility varies widely across

different taxa (Mudge et al., 2009). Four potential mechanisms of interspecific

incompatibility are identified by Andrews and Marquez (1993): cellular recognition,

wounding response, plant growth regulators, and incompatibility toxins. If the 3205 scion

experienced a degree of incompatibility through any of these mechanisms that the 3212

scion did not, it may have resulted in the comparatively stunted plant growth observed in

this experiment.

72

Average internode length was greater for every genotype in control conditions

compared to hot conditions, and for 3205 and 3205xS.torvum the difference was

significant (Table 3). Internode length of tomatoes decreases in heat stress (Wahid et

al., 2007), but since both scion genotypes experienced the same internode length

reduction as their non-grafted controls, we can not determine whether grafting onto S.

torvum has any effect on tomato internode length when grown under heat stress

conditions.

The analysis of stomatal resistances among genotypes demonstrates increased

heat tolerance when S. torvum is utilized as a rootstock. For both cultivars, average

stomatal resistance was significantly reduced in hot conditions for the non-grafted

controls while there was an insignificant reduction when grafted onto S. torvum rootstock

(Table 4). Stomatal resistance is known to increase when tomatoes are exposed to

drought stress conditions in order to conserve water, but will decrease as temperatures

rise, as the plant intakes more carbon dioxide to accommodate increased respiration

(Camejo et al., 2005; Sobeih et al., 2004). Diligent irrigation ensured there was no

drought stress in this experiment. Thus, since the drop in stomatal resistance was

significantly more in the non-grafted controls than the grafted genotypes, we concluded

the grafted genotypes were not reacting to the same extent as the heat stress

treatments. The lower resistances of the non-grafted plants in heat stress may also have

contributed to the observed height increase when compared to grafted plants in hot

conditions (Table 2). As mentioned before, the lower resistances likely accommodated

higher respiration rates, which could have led to a faster rate of growth.

Increasing temperatures worldwide make the development of heat-tolerant plants

and cultural practices a pressing need. The insignificant drop in stomatal resistance of

both tomato scions tested, CLN 3205A and CLN 3212A, when grafted onto S. torvum

73

imply that they do not react as strongly to heat stress compared to non-grafted controls

of the same genotype. Further research needs to be conducted to determine if this

observed stomatal stability would result in increased flower bud initiation, fruit set, and

ultimately higher yields.

74


Abdalla, A.A. and K. Verderk. 1968. Growth, flowering and fruit set of tomato at high temperature. The Neth J Agric Sci 16:71-76.

Abdelmageed, A. and N. Gruda. 2009. Influence of grafting on growth, development and some physiological parameters of tomatoes under controlled heat stress conditions. European Journal of Horticultural Science 74(1):16-20.

Abdul-Baki, A.A. 1991. Tolerance of tomato cultivars and selected germplasm to heat stress. J. Am. Soc. Hort. Sci. 116(6):1113-1116.

Ahn, S., Y. Im, G. Chung, B. Cho, and S. Suh. 1999. Physiological responses of grafted-cucumber leaves and rootstock roots affected by low root temperature. Scientia horticulturae 81(4):397-408.







Bulder, H., P.R. Van Hasselt, and P. Kuiper. 1987. The effect of temperature on early growth of cucumber genotypes differing in genetic adaptation to low-energy conditions. Scientia horticulturae 31(1):53-60.




Foolad, M. and G. Lin. 2001. Genetic analysis of cold tolerance during vegetative growth in tomato, Lycopersicon esculentum Mill. Euphytica 122(1):105-111.

75










Miller, S.A., A.N. Karim, M. Razaul, A.M. Baltazar, E.G. Rajotte, and G. Norton. 2005. Developing IPM packages in Asia. p. 27-50. In: G.W. Norton, E.A. Heinrichs, G.C. Luther and M.E. Irwin (eds.). Globalizing Integrated Pest Management – A Participatory ResearchProcess. Blackwell Publishing, Oxford.




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76








Tachibana, S. 1982. Comparison of effects of root temperature on the growth and mineral nutrition of cucumber cultivars and figleaf gourd. Journal of the Japanese Society for Horticultural Science 51.


Van der Ploeg, A. and E. Heuvelink. 2005. Influence of sub-optimal temperature on tomato growth and yield. Journal of horticultural science & biotechnology 80(6):652-659.


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77

Table 1. Scion and rootstock combinations were analyzed for morphological and physiological responses to different environmental pressures.

Scion Rootstock Final 'Genotype'

# Replications per Environmental Treatment


3

CLN 3205A Solanum torvum "3205xS. torvum" 3


3

CLN 3212A Solanum torvum "3212xS. torvum" 3

Table 2. Average final plant height of all genotypes in each environmental condition. Letters denote statistical differences (p<0.05) within rows. Stars denote statistical differences (p<0.05) within columns.

Plant Height (cm) Genotype

3205 3205xS.torvum 3212 3212xS.torvum (n) 3 3 3 3 Control 72 a 64 ac 46 b 58 c * * Hot 60 52.33 57 52.67

Table 3. Average final internode length of all genotypes in each environmental condition. Stars denote statistical significance (p<0.05) within columns.

Internode Length (cm)

Genotype 3205 3205xS.torvum 3212 3212xS.torvum

(n) 3 3 3 3 Control 5.7 6.22 4.83 4.73 * * Hot 4.39 4.5 4 4.66

78

Table 4. Average final stomatal resistance of all genotypes in each environmental condition. Letters denote statistical differences (p<0.05) within rows. Stars denote statistical significance (p<0.05) within columns.

Stomatal Resistance

(mmol/m2/s)

Genotype

3205 3205xS.torvum 3212 3212xS.torvum

(n) 3 3 3 3 Control 2.26 1.09 1.42 1.08 * * Hot 0.24 0.32 0.38 0.52

79

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87

APPENDIX

Data Tables for ANOVA and Regression Analysis

88

Table A1: Rootstock Compatibility Trials

Scion Rootstock Replication Days to Fusion Survival 3212 non-‐grafted 1 0 1 3212 non-‐grafted 2 0 1 3212 non-‐grafted 3 0 1 3212 non-‐grafted 4 0 1 3212 non-‐grafted 5 0 1 3212 non-‐grafted 6 0 1 3212 non-‐grafted 7 0 1 3212 non-‐grafted 8 0 1 3212 non-‐grafted 9 0 1 3212 non-‐grafted 10 0 1 3212 3212 1

0

3212 3212 2 11 1 3212 3212 3 11 1 3212 3212 4 11 1 3212 3212 5 11 1 3212 3212 6 11 1 3212 3212 7

0

3212 3212 8 11 1 3212 3212 9 11 1 3212 3212 10 11 1 3212 Maxifort 1 8 1 3212 Maxifort 2 8 1 3212 Maxifort 3 8 1 3212 Maxifort 4 8 1 3212 Maxifort 5 8 1 3212 Maxifort 6 10 1 3212 Maxifort 7 10 1 3212 Maxifort 8 13 1 3212 Maxifort 9 13 1 3212 Maxifort 10 13 1 3212 S. torvum 1 10 1 3212 S. torvum 2 10 1 3212 S. torvum 3 10 1 3212 S. torvum 4 10 1 3212 S. torvum 5 10 1 3212 S. torvum 6 14 1 3212 S. torvum 7 13 1 3212 S. torvum 8

0

3212 S. torvum 9 13 1 3212 S. torvum 10 11 0

89

Celebrity non-‐grafted 1 0 1 Celebrity non-‐grafted 2 0 1 Celebrity non-‐grafted 3 0 1 Celebrity non-‐grafted 4 0 1 Celebrity non-‐grafted 5 0 1 Celebrity non-‐grafted 6 0 1 Celebrity non-‐grafted 7 0 1 Celebrity non-‐grafted 8 0 1 Celebrity non-‐grafted 9 0 1 Celebrity non-‐grafted 10 0 1 Celebrity Celebrity 1 11 1 Celebrity Celebrity 2 11 1 Celebrity Celebrity 3 11 1 Celebrity Celebrity 4 19 1 Celebrity Celebrity 5

0

Celebrity Celebrity 6 11 1 Celebrity Celebrity 7

0

Celebrity Celebrity 8 11 1 Celebrity Celebrity 9 11 1 Celebrity Celebrity 10

0

Celebrity Maxifort 1 8 1 Celebrity Maxifort 2 8 1 Celebrity Maxifort 3 8 1 Celebrity Maxifort 4 8 1 Celebrity Maxifort 5 8 1 Celebrity Maxifort 6 13 1 Celebrity Maxifort 7 13 1 Celebrity Maxifort 8 13 1 Celebrity Maxifort 9 13 1 Celebrity Maxifort 10 13 1 Celebrity S. torvum 1 10 1 Celebrity S. torvum 2 14 1 Celebrity S. torvum 3 10 1 Celebrity S. torvum 4 10 1 Celebrity S. torvum 5 10 1 Celebrity S. torvum 6 14 1 Celebrity S. torvum 7 10 1 Celebrity S. torvum 8 15 1 Celebrity S. torvum 9 15 1 Celebrity S. torvum 10 15 1

3212 S. torvum Veg 1

0 3212 S. torvum Veg 2 12 1 3212 S. torvum Veg 3

0

90

3212 S. torvum Veg 4 12 1 Celebrity S. torvum Veg 1

0

Celebrity S. torvum Veg 2

0 Celebrity S. torvum Veg 3 12 1 Celebrity S. torvum Veg 4 12 1

91

Table A2: Moisture Condition Trials

Genotype Day Replication Treatment Resistance Height Nodes 3212 1 1 wet 0.95 30 7 3212 1 2 wet 0.98 43 10 3212 1 3 wet 0.46 52 9

3212x3212 1 1 wet 1.81 61 9 3212x3212 1 2 wet 1.3 34 8 3212x3212 1 3 wet 1.73 50 9 3212xS.torvum 1 1 wet 1.33 50 10 3212xS.torvum 1 2 wet 2.66 51 11 3212xS.torvum 1 3 wet 1.24 49 11 3212xMaxifort 1 1 wet 0.66 56 10 3212xMaxifort 1 2 wet 0.94 57 10 3212xMaxifort 1 3 wet 0.42 65 9 Celebrity 1 1 wet 0.92 50 10 Celebrity 1 2 wet 0.66 44 10 Celebrity 1 3 wet 1.86 54 10 CelebrityxCelebrity 1 1 wet 1.11 40 7 CelebrityxCelebrity 1 2 wet 1.34 42 8 CelebrityxCelebrity 1 3 wet 0.93 49 10 CelebrityxS.torvum 1 1 wet 11.8 18 4 CelebrityxS.torvum 1 2 wet 1.04 33 7 CelebrityxS.torvum 1 3 wet 1.01 26 7 CelebrityxMaxifort 1 1 wet 1.36 51 11 CelebrityxMaxifort 1 2 wet 1.22 49 11 CelebrityxMaxifort 1 3 wet 1.3 45 10

3212 1 1 dry 1.27 39 10 3212 1 2 dry 1.2 57 10 3212 1 3 dry 1.07 26 7

3212x3212 1 1 dry 0.74 31 7 3212x3212 1 2 dry 0.47 58 11 3212x3212 1 3 dry 1.05 36 8 3212xS.torvum 1 1 dry 0.62 51 10 3212xS.torvum 1 2 dry 0.83 42 8 3212xMaxifort 1 1 dry 0.98 46 9 3212xMaxifort 1 2 dry 0.93 52 9 3212xMaxifort 1 3 dry 0.9 46 10 Celebrity 1 1 dry 0.68 47 9 Celebrity 1 2 dry 1.65 43 9 Celebrity 1 3 dry 1.14 47 11 CelebrityxCelebrity 1 1 dry 1.53 47 10 CelebrityxCelebrity 1 2 dry 6.2 25 4 CelebrityxCelebrity 1 3 dry 2.18 43 8 CelebrityxS.torvum 1 1 dry 1.14 22 6 CelebrityxS.torvum 1 2 dry 3.75 24 6

92

CelebrityxS.torvum 1 3 dry 3.15 25 6 CelebrityxMaxifort 1 1 dry 2 37 10 CelebrityxMaxifort 1 2 dry 2.66 49 12 CelebrityxMaxifort 1 3 dry 1.82 38 9

3212 1 1 optimal 2.64 31 7 3212x3212 1 1 optimal 2.22 46 9 3212xS.torvum 1 1 optimal 4.9 41 11 3212xS.torvum 1 2 optimal 1.5 32 6 3212xS.torvum 1 3 optimal 1.4 48 10 3212xMaxifort 1 1 optimal 1.67 52 9 3212xMaxifort 1 2 optimal 1.18 43 9 3212xMaxifort 1 3 optimal 1.61 32 8 Celebrity 1 1 optimal 0.77 46 12 Celebrity 1 2 optimal 1.57 45 11 Celebrity 1 3 optimal 1.52 41 12 CelebrityxCelebrity 1 1 optimal 2.04 33 8 CelebrityxCelebrity 1 2 optimal 1.68 40 9 CelebrityxS.torvum 1 1 optimal 1.37 38 7 CelebrityxS.torvum 1 2 optimal 2.2 38 9 CelebrityxS.torvum 1 3 optimal 1.36 25 8 CelebrityxMaxifort 1 1 optimal 1.5 44 10 CelebrityxMaxifort 1 2 optimal 1.22 46 10 CelebrityxMaxifort 1 3 optimal 0.8 44 12

3212 5 1 wet 0.68 40 8 3212 5 2 wet 0.83 49 10 3212 5 3 wet 1.25 59 11

3212x3212 5 1 wet 1.77 72 8 3212x3212 5 2 wet 1.65 43 9 3212x3212 5 3 wet 0.91 58 9 3212xS.torvum 5 1 wet 1.16 57 11 3212xS.torvum 5 2 wet 5 58 12 3212xS.torvum 5 3 wet 0.82 59 13 3212xMaxifort 5 1 wet 0.65 68 12 3212xMaxifort 5 2 wet 1.55 72 12 3212xMaxifort 5 3 wet 0.96 72 9 Celebrity 5 1 wet 0.92 58 11 Celebrity 5 2 wet 0.5 52 12 Celebrity 5 3 wet 1.21 57 13 CelebrityxCelebrity 5 1 wet 1.21 50 9 CelebrityxCelebrity 5 2 wet 0.7 54 11 CelebrityxCelebrity 5 3 wet 0.7 61 12 CelebrityxS.torvum 5 1 wet 5.4 17 5 CelebrityxS.torvum 5 2 wet 1.28 41 11 CelebrityxS.torvum 5 3 wet 1.92 31 9 CelebrityxMaxifort 5 1 wet 1.24 59 10 CelebrityxMaxifort 5 2 wet 1.09 55 13

93

CelebrityxMaxifort 5 3 wet 1.36 53 12 3212 5 1 dry 10.2 50 9 3212 5 2 dry 1.61 62 11 3212 5 3 dry 8.1 37 8

3212x3212 5 1 dry 2.36 40 9 3212x3212 5 2 dry 2.78 65 11 3212x3212 5 3 dry 0.85 45 10 3212xS.torvum 5 1 dry 7.9 61 11 3212xS.torvum 5 2 dry 6.3 48 9 3212xMaxifort 5 1 dry 16.3 52 11 3212xMaxifort 5 2 dry 1.25 56 11 3212xMaxifort 5 3 dry 6.3 50 9 Celebrity 5 1 dry 6.9 56 11 Celebrity 5 2 dry 4.85 50 13 Celebrity 5 3 dry 5.8 56 13 CelebrityxCelebrity 5 1 dry 13.8 51 11 CelebrityxCelebrity 5 2 dry 5.7 25 5 CelebrityxCelebrity 5 3 dry 12.9 48 9 CelebrityxS.torvum 5 1 dry 1.49 24 8 CelebrityxS.torvum 5 2 dry 1.34 28 8 CelebrityxS.torvum 5 3 dry 1.97 30 8 CelebrityxMaxifort 5 1 dry 1.63 47 9 CelebrityxMaxifort 5 2 dry 7.7 56 13 CelebrityxMaxifort 5 3 dry 2.42 45 10


3212 10 1 wet 1.08 56 13 3212 10 2 wet 0.46 59 13 3212 10 3 wet 0.61 71 12

94


3212 10 1 dry 8.2 53 11 3212 10 2 dry 6.7 63 11 3212 10 3 dry 10.4 40 9

3212x3212 10 1 dry 3.65 52 10 3212x3212 10 2 dry 9.6 59 14 3212x3212 10 3 dry 4.75 53 12 3212xS.torvum 10 1 dry 4.25 63 11 3212xS.torvum 10 2 dry 1.55 59 10 3212xMaxifort 10 1 dry 2.62 52 11 3212xMaxifort 10 2 dry 6.1 55 11 3212xMaxifort 10 3 dry 9 56 12 Celebrity 10 1 dry 6.2 57 13 Celebrity 10 2 dry 3.8 54 15 Celebrity 10 3 dry 7.6 57 15 CelebrityxCelebrity 10 1 dry 7.9 53 14 CelebrityxCelebrity 10 2 dry 1.83 26 5 CelebrityxCelebrity 10 3 dry 6.3 57 10 CelebrityxS.torvum 10 1 dry 1.38 30 10 CelebrityxS.torvum 10 2 dry 1.47 32 9 CelebrityxS.torvum 10 3 dry 1.16 32 10 CelebrityxMaxifort 10 1 dry 4.5 49 12 CelebrityxMaxifort 10 2 dry 6.8 57 12 CelebrityxMaxifort 10 3 dry 6.4 49 11

3212 10 1 optimal 0.69 55 12 3212x3212 10 1 optimal 0.8 70 14

95

3212xS.torvum 10 1 optimal 1.15 50 10 3212xS.torvum 10 2 optimal 0.77 59 10 3212xS.torvum 10 3 optimal 1.43 64 11 3212xMaxifort 10 1 optimal 0.97 64 12 3212xMaxifort 10 2 optimal 0.76 65 14 3212xMaxifort 10 3 optimal 0.88 57 13 Celebrity 10 1 optimal 0.73 67 17 Celebrity 10 2 optimal 0.63 63 14 Celebrity 10 3 optimal 0.33 59 12 CelebrityxCelebrity 10 1 optimal 0.46 54 12 CelebrityxCelebrity 10 2 optimal 0.67 61 12 CelebrityxS.torvum 10 1 optimal 0.73 55 12 CelebrityxS.torvum 10 2 optimal 0.58 53 12 CelebrityxS.torvum 10 3 optimal 0.62 34 10 CelebrityxMaxifort 10 1 optimal 0.45 60 12 CelebrityxMaxifort 10 2 optimal 0.49 64 13 CelebrityxMaxifort 10 3 optimal 0.68 64 13

3212 15 1 wet 0.72 79 15 3212 15 2 wet 0.46 72 14 3212 15 3 wet 1.39 67 13


3212 15 1 dry 3.85 52 10 3212 15 2 dry 3.4 65 11 3212 15 3 dry 10.1 41 9

3212x3212 15 1 dry 18.7 58 10 3212x3212 15 2 dry 4.5 66 9

96

3212x3212 15 3 dry 8 55 10 3212xS.torvum 15 1 dry 5.1 73 10 3212xS.torvum 15 2 dry 2.62 50 12 3212xMaxifort 15 1 dry 6.2 56 12 3212xMaxifort 15 2 dry 15.9 59 11 3212xMaxifort 15 3 dry 25.6 49 12 Celebrity 15 1 dry 12 55 13 Celebrity 15 2 dry 15.3 51 13 Celebrity 15 3 dry 14.2 53 13 CelebrityxCelebrity 15 1 dry 14.5 48 12 CelebrityxCelebrity 15 2 dry 0.73 27 6 CelebrityxCelebrity 15 3 dry 3.65 47 11 CelebrityxS.torvum 15 1 dry 6.9 35 10 CelebrityxS.torvum 15 2 dry 4.8 38 11 CelebrityxS.torvum 15 3 dry 4.15 36 11 CelebrityxMaxifort 15 1 dry 11 42 11 CelebrityxMaxifort 15 2 dry 5.1 53 12 CelebrityxMaxifort 15 3 dry 13.1 50 13


3212 20 1 wet 1.67 73 12 3212 20 2 wet 0.91 81 12 3212 20 3 wet 2.08 83 14

3212x3212 20 1 wet 3.15 81 12 3212x3212 20 2 wet 1.52 82 12 3212x3212 20 3 wet 1.23 79 13 3212xS.torvum 20 1 wet 1.52 80 15 3212xS.torvum 20 2 wet 3.8 80 15 3212xS.torvum 20 3 wet 2.36 92 16

97

3212xMaxifort 20 1 wet 1.85 83 15 3212xMaxifort 20 2 wet 1.52 94 16 3212xMaxifort 20 3 wet 1.5 94 13 Celebrity 20 1 wet 1.39 94 16 Celebrity 20 2 wet 0.82 85 13 Celebrity 20 3 wet 1.5 86 14 CelebrityxCelebrity 20 1 wet 0.78 78 13 CelebrityxCelebrity 20 2 wet 2.26 86 16 CelebrityxCelebrity 20 3 wet 1.7 89 14 CelebrityxS.torvum 20 1 wet 1.36 32 10 CelebrityxS.torvum 20 2 wet 1.59 68 11 CelebrityxS.torvum 20 3 wet 0.77 45 11 CelebrityxMaxifort 20 1 wet 1.52 83 13 CelebrityxMaxifort 20 2 wet 1.17 73 14 CelebrityxMaxifort 20 3 wet 1.4 75 17

3212 20 1 dry 12 50 11 3212 20 2 dry 0 57 12 3212 20 3 dry 11 40 9

3212x3212 20 1 dry 19.3 51 11 3212x3212 20 2 dry 0 61 12 3212x3212 20 3 dry 0 46 11 3212xS.torvum 20 1 dry 15.6 70 13 3212xS.torvum 20 2 dry 18 50 11 3212xMaxifort 20 1 dry 39 53 13 3212xMaxifort 20 2 dry 0 57 12 3212xMaxifort 20 3 dry 0 47 11 Celebrity 20 1 dry 20.2 53 11 Celebrity 20 2 dry 19.8 53 12 Celebrity 20 3 dry 23.6 52 13 CelebrityxCelebrity 20 1 dry 0 43 7 CelebrityxCelebrity 20 2 dry 2.48 33 12 CelebrityxCelebrity 20 3 dry 25.6 47 11 CelebrityxS.torvum 20 1 dry 12.5 33 10 CelebrityxS.torvum 20 2 dry 9.8 36 10 CelebrityxS.torvum 20 3 dry 17 36 12 CelebrityxMaxifort 20 1 dry 18.4 40 12 CelebrityxMaxifort 20 2 dry 0 53 13 CelebrityxMaxifort 20 3 dry 19.2 41 12

3212 20 1 optimal 1.94 77 13 3212x3212 20 1 optimal 2.08 90 16 3212xS.torvum 20 1 optimal 2.44 66 12 3212xS.torvum 20 2 optimal 2.5 62 12 3212xS.torvum 20 3 optimal 2.22 77 14 3212xMaxifort 20 1 optimal 1.24 79 14 3212xMaxifort 20 2 optimal 1.43 81 14 3212xMaxifort 20 3 optimal 1.12 77 14

98

Celebrity 20 1 optimal 1.45 83 14 Celebrity 20 2 optimal 1.3 75 14 Celebrity 20 3 optimal 1.62 76 13 CelebrityxCelebrity 20 1 optimal 0.88 70 14 CelebrityxCelebrity 20 2 optimal 2 63 14 CelebrityxS.torvum 20 1 optimal 0.92 70 13 CelebrityxS.torvum 20 2 optimal 0.91 63 13 CelebrityxS.torvum 20 3 optimal 0.63 55 13 CelebrityxMaxifort 20 1 optimal 1.31 77 13 CelebrityxMaxifort 20 2 optimal 1.56 82 13 CelebrityxMaxifort 20 3 optimal 1.41 77 15

3212 25 1 wet 1.38 85 13 3212 25 2 wet 1.03 85 13 3212 25 3 wet 1.25 73 14


3212 25 1 dry 0 50 11 3212 25 2 dry 0 57 12 3212 25 3 dry 0 40 9

3212x3212 25 1 dry 0 51 11 3212x3212 25 2 dry 0 61 12 3212x3212 25 3 dry 0 46 11 3212xS.torvum 25 1 dry 0 70 13 3212xS.torvum 25 2 dry 0 50 11 3212xMaxifort 25 1 dry 0 53 13 3212xMaxifort 25 2 dry 0 57 12 3212xMaxifort 25 3 dry 0 47 11

99

Celebrity 25 1 dry 0 53 11 Celebrity 25 2 dry 0 53 12 Celebrity 25 3 dry 0 52 13 CelebrityxCelebrity 25 1 dry 0 43 7 CelebrityxCelebrity 25 2 dry 0 33 12 CelebrityxCelebrity 25 3 dry 0 47 11 CelebrityxS.torvum 25 1 dry 0 33 10 CelebrityxS.torvum 25 2 dry 0 36 10 CelebrityxS.torvum 25 3 dry 0 36 12 CelebrityxMaxifort 25 1 dry 0 40 12 CelebrityxMaxifort 25 2 dry 0 53 13 CelebrityxMaxifort 25 3 dry 0 41 12


100

Table A3: Moisture Trial Final Day Calculations

Genotype Replication Treatment Fresh Weight Dry Weight Leaf Area 3212 1 wet 198.71 38.47 30.78 3212 2 wet 202.37 42.8 33.62 3212 3 wet 102.42 23.59 15.96

3212x3212 1 wet 192.3 37.2 23.87 3212x3212 2 wet 199.4 30.24 24.75 3212x3212 3 wet 201.52 38.06 22.42 3212xS.torvum 1 wet 172.8 41.02 27.75 3212xS.torvum 2 wet 188.7 34.6 23.31 3212xS.torvum 3 wet 214.36 43.58 30.93 3212xMaxifort 1 wet 238.98 42.6 28.16 3212xMaxifort 2 wet 238.2 48.05 24.99 3212xMaxifort 3 wet 211.75 36.6 36.84 Celebrity 1 wet 190.01 43.4 43.36 Celebrity 2 wet 177.36 38.31 32.85 Celebrity 3 wet 192.63 43.85

CelebrityxCelebrity 1 wet 217.91 37.77 CelebrityxCelebrity 2 wet 175.06 37.22 38.74

CelebrityxCelebrity 3 wet 186.86 41.8 34.99 CelebrityxS.torvum 1 wet 79.87 6.54 22.09 CelebrityxS.torvum 2 wet 115.85 23.78 24.49 CelebrityxS.torvum 3 wet 152.3 23.35 25.69 CelebrityxMaxifort 1 wet 240.6 45.95

CelebrityxMaxifort 2 wet 244.73 50.55 20.46 CelebrityxMaxifort 3 wet 201.72 42.57 29.56

3212 1 dry 9.88 8.98 6.67 3212 2 dry 10.57 9.16 4.63 3212 3 dry 7.41 6.93 5.08

3212x3212 1 dry 6.68 6.19 10.06 3212x3212 2 dry 9.02 8.31 4.62 3212x3212 3 dry 6.55

4.02

3212xS.torvum 1 dry 11.12 9.81 8.15 3212xS.torvum 2 dry 7.48 6.25 4.09 3212xMaxifort 1 dry 5.7 4.95 4.02 3212xMaxifort 2 dry 7.59 6.42 5.51 3212xMaxifort 3 dry 8.34

5.7

Celebrity 1 dry 8.28 7.4 9.89 Celebrity 2 dry 8.08 7.48 4.7 Celebrity 3 dry 8.37 7.54 4.12 CelebrityxCelebrity 1 dry 10.45 8.98 3.13 CelebrityxCelebrity 2 dry 2.95 2.69 2.23 CelebrityxCelebrity 3 dry 7.74 7.21 5.04 CelebrityxS.torvum 1 dry 3.48 2.9 3 CelebrityxS.torvum 2 dry 4.51 4.29 3.05

101

CelebrityxS.torvum 3 dry 5.41 5.01 4.02 CelebrityxMaxifort 1 dry 7.13

4.86

CelebrityxMaxifort 2 dry 7.93 7.5 5.94 CelebrityxMaxifort 3 dry 5.63 5.25 3.75

3212 1 optimal 202.05 28.31 18.57 3212x3212 1 optimal 238.48 44.19 33.65 3212xS.torvum 1 optimal 236.55 39.44 39.62 3212xS.torvum 2 optimal 286.53 42.55 20.32 3212xS.torvum 3 optimal 292.78 47.94 27.84 3212xMaxifort 1 optimal 245.55 51.12 25.82 3212xMaxifort 2 optimal 279.9 59.14 28.21 3212xMaxifort 3 optimal 288.43 56.85 38.11 Celebrity 1 optimal 304.11 63.98 28.86 Celebrity 2 optimal 276.15 62.1 37 Celebrity 3 optimal 230.32 56.51 10.47 CelebrityxCelebrity 1 optimal 304.94 59.52 20.7 CelebrityxCelebrity 2 optimal 276.12 59.3 35.08 CelebrityxS.torvum 1 optimal 570 67.23 36.21 CelebrityxS.torvum 2 optimal 450 56.07 25.44 CelebrityxS.torvum 3 optimal 522 62.75 37.51 CelebrityxMaxifort 1 optimal 484 70.03 49.09 CelebrityxMaxifort 2 optimal 487 66.21 42.73 CelebrityxMaxifort 3 optimal 467 59.5 41.11

102

Table A4: Heat Condition Trials

Genotype Day Replication Treatment Resistance Height Nodes 3205 1 1 control 3.1 19 4 3205 1 2 control 3.55 18 5 3205 1 3 control 3.35 22 5 3205 1 4 hot 5.7 23 5 3205 1 5 hot 11.3 25 8 3205 1 6 hot 3.65 22 6 3212 1 1 control 1.82 15 5 3212 1 2 control 1.33 18 5 3212 1 3 control 1.53 17 5 3212 1 4 hot 2.04 17 6 3212 1 5 hot 1.2 16 5 3212 1 6 hot 1.27 14 5

3205xS.torvum 1 1 control 4.7 21 5 3205xS.torvum 1 2 control 1.68 21 5 3205xS.torvum 1 3 control 1.85 17 5 3205xS.torvum 1 4 hot 3.95 22 4 3205xS.torvum 1 5 hot 1.57 22 5 3205xS.torvum 1 6 hot 1.8 25 7 3212xS.torvum 1 1 control 1.78 20 5 3212xS.torvum 1 2 control 2.02 26 8 3212xS.torvum 1 3 control 2.8 19 5 3212xS.torvum 1 4 hot 1.27 20 6 3212xS.torvum 1 5 hot 1.88 18 5 3212xS.torvum 1 6 hot 2.02 18 5

3205 5 1 control 0.68 26 8 3205 5 2 control 0.67 23 8 3205 5 3 control 0.46 29 8 3205 5 4 hot 0.83 30 9 3205 5 5 hot 0.76 32 10 3205 5 6 hot 1 30 9 3212 5 1 control 0.6 21 7 3212 5 2 control 0.64 23 7 3212 5 3 control 0.5 23 7 3212 5 4 hot 0.65 26 9 3212 5 5 hot 0.48 25 7 3212 5 6 hot 0.75 21 7

3205xS.torvum 5 1 control 0.66 21 6 3205xS.torvum 5 2 control 0.63 20 6 3205xS.torvum 5 3 control 0.63 21 8 3205xS.torvum 5 4 hot 1 26 7 3205xS.torvum 5 5 hot 1 28 8 3205xS.torvum 5 6 hot 1 25 8 3212xS.torvum 5 1 control 0.86 22 6

103

3212xS.torvum 5 2 control 0.87 28 7 3212xS.torvum 5 3 control 0.81 23 6 3212xS.torvum 5 4 hot 0.71 26 9 3212xS.torvum 5 5 hot 0.8 26 7 3212xS.torvum 5 6 hot 0.92 21 7

3205 15 1 control 0.53 49 11 3205 15 2 control 0.49 43.5 11 3205 15 3 control 0.35 51 10 3205 15 4 hot 0.37 41 12 3205 15 5 hot 1.1 44 13 3205 15 6 hot 0.35 42 11 3212 15 1 control 0.15 39 11 3212 15 2 control 0.14 45 8 3212 15 3 control 0.24 39 8 3212 15 4 hot 0.035 45 12 3212 15 5 hot 0.28 34 9 3212 15 6 hot 0.25 35 9


3205 25 1 control 0.66 68 13 3205 25 2 control 0.67 64 12 3205 25 3 control 0.78 61 12 3205 25 4 hot 0.24 53 15 3205 25 5 hot 0.25 47 14 3205 25 6 hot 0.62 54 15 3212 25 1 control 0.49 60 13 3212 25 2 control 2.04 59 11 3212 25 3 control 2.52 57 12 3212 25 4 hot 0.36 54 13 3212 25 5 hot 0.33 47 12 3212 25 6 hot 0.35 48 11

3205xS.torvum 25 1 control 0.81 45 11 3205xS.torvum 25 2 control 0.37 35 10 3205xS.torvum 25 3 control 0.78 42 10 3205xS.torvum 25 4 hot 0.45 45 12 3205xS.torvum 25 5 hot 1.19 59 15

104

3205xS.torvum 25 6 hot 0.61 51 13 3212xS.torvum 25 1 control 0.69 50 11 3212xS.torvum 25 2 control 0.97 55 12 3212xS.torvum 25 3 control 1.24 50 12 3212xS.torvum 25 4 hot 1.14 55 13 3212xS.torvum 25 5 hot 0.34 50 12 3212xS.torvum 25 6 hot 1.25 44 10

3205 35 1 control 1.7 75 14 3205 35 2 control 2.32 71 12 3205 35 3 control 2.78 70 12 3205 35 4 hot 0.31 59 13 3205 35 5 hot 0.21 55 14 3205 35 6 hot 0.2 66 14 3212 35 1 control 1.42 52 9 3212 35 2 control 1.39 42 11 3212 35 3 control 1.45 44 9 3212 35 4 hot 0.38 50 12 3212 35 5 hot 0.46 60 16 3212 35 6 hot 0.29 61 15

Documents

Interspecific Grafting of Tomato (Solanum lycopersicum